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Cytogenetics in acute leukemia
Blood Reviews (2004) 18, 115–136
www.elsevierhealth.com/journals/blre
Cytogenetics in acute leukemia zeka,*, Nyla A. Heeremab, Clara D. Bloomfielda Krzysztof Mro a
Division of Hematology and Oncology, The Comprehensive Cancer Center, The Arthur G. James Cancer Hospital and Richard J. Solove Research Institute, Room 1248B, The Ohio State University, 300 West Tenth Avenue, Columbus, OH 43210-1228, USA b Department of Pathology, The Comprehensive Cancer Center, The Ohio State University, Columbus, OH 43210-1228, USA
KEYWORDS Acute myeloid leukemia; Acute lymphoblastic leukemia; Chromosome aberrations; Human; Karyotyping; Prognosis
Summary Cytogenetic analyses in acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) have revealed a great number of non-random chromosome abnormalities. In many instances, molecular studies of these abnormalities identified specific genes implicated in the process of leukemogenesis. The more common chromosome aberrations have been associated with specific laboratory and clinical characteristics, and are now being used as diagnostic and prognostic markers guiding the clinician in selecting the most effective therapies. Specific chromosome aberrations and their molecular counterparts have been included in the World Health Organization classification of hematologic malignancies, and together with morphology, immunophenotype and clinical features are used to define distinct disease entities. However, the prognostic importance of less frequent recurrent aberrations in AML and ALL, both primary and secondary, is still to be determined. This review summarizes current views on clinical relevance of major cytogenetic findings in adult AML and ALL. c 2003 Elsevier Ltd. All rights reserved.
Introduction The role of cytogenetics in determining the biologic basis of acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) is now widely recognized. By identifying acquired chromosome aberrations that recur in AML and ALL, and providing precise chromosomal location of breakpoints in leukemia-associated translocations and inversions, cytogenetics aided in cloning of many genes whose activation or fusion with other genes contributes to the neoplastic process.1–3 Further
*
Corresponding author. Tel.: +1-614-293-3150; fax: +1-614293-3457. zek). E-mail address: [email protected] (K. Mro
characterization of these genes revealed that they are often involved directly or indirectly in the development and homeostasis of normal blood cells, and that abnormal protein products of fusion genes created by specific translocations and inversions can dysregulate proliferation, differentiation or programmed cell death (apoptosis) of blood cell precursors.1 This has paved the way to designing novel therapeutic agents targeting specific genetic abnormalities in leukemic blasts, such as imatinib mesylate, the Bcr–Abl tyrosine kinase inhibitor that suppresses proliferation of cells harboring the BCR–ABL fusion gene created by t(9;22) (q34;q11.2), a recurrent chromosome aberration in chronic myelogenous leukemia (CML) and ALL.4 Furthermore, cytogenetic analysis is now routine part of clinical practice, being an important
0268-960X/$ - see front matter c 2003 Elsevier Ltd. All rights reserved. doi:10.1016/S0268-960X(03)00040-7
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tool in diagnosis and prognostication of acute leukemias. Specific chromosome aberrations and their molecular counterparts have been included in the World Health Organization (WHO) classification of hematologic malignancies, and together with morphology, immunophenotype and clinical features are being used to define distinct disease entities.5–7 Pretreatment cytogenetic findings have been repeatedly shown to be among the most important, independent prognostic factors in both AML8–20 and ALL.21–26 Consequently, cytogenetic analyses are considered mandatory for analyzing outcome of many clinical trials and are currently used to stratify patients for different types of therapy.27–29 In this review, we will summarize the clinical relevance of major cytogenetic findings in adult AML and ALL.
Acute myeloid leukemia Clonal chromosome abnormalities, that is, an identical structural aberration or gain of the same, structurally intact chromosome detected in at least two metaphase cells or the same chromosome missing from a minimum of three cells, are consistently found in the majority of AML patients at diagnosis. However, in contrast to patients diagnosed with CML, who are invariably positive for t(9;22) or its variants, the cytogenetic picture of AML is much more complex. To date, approximately 200 different structural and numerical aberrations such as reciprocal translocations, inversions, insertions, deletions, unbalanced translocations, isochromosomes, isodicentric chromosomes, isolated trisomies and monosomies have been found to be recurring chromosome changes in AML. Many of these aberrations are very rare, being so far detected in a few patients worldwide, whereas others occur more frequently. These more common abnormalities, together with their frequencies among adults and children with AML, are presented in Table 1. A more complete list of specific AML-associated recurrent chromosome aberrations, and genes affected by their formation (if known), can be found in Refs. 27, 30–32. The incidence of abnormal karyotypes is in general lower in adult than in pediatric de novo AML. While 55% of 4257 adults with AML enrolled onto three large cooperative studies displayed chromosome aberrations (range 53–60%), 76% of 1184 patients included in the four largest series of childhood AML had an abnormal karyotype (range 68–85%; Table 1). The reasons for such a discrep-
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ancy are unknown, but may be related to biological differences between pediatric and adult disease. This is likely reflected by varying proportions of specific chromosome aberrations in children and adults with AML. For example, balanced rearrangements, mostly reciprocal translocations, involving band 11q23 and disrupting the MLL gene are on average four times more common in children than in adults. The frequency of 11q23 rearrangements in patients with AML decreases appreciably with age: they are detected in 43–58% of infants aged 12 months or less,33–35 in 39% of children aged 13–24 months,35 in 8–9% of children older than 24 months,35 and in just 4–7% of adults (Table 1), among whom, according to the United Kingdom Medical Research Council (MRC) study,19 the frequency of 11q23 rearrangements goes down from 5% in patients aged 15–34 years to 2% among those aged 35–55 years and 1% in patients older than 55 years. Some other, relatively rare, recurrent aberrations are essentially restricted to young children with AML, not being detected in adult patients thus far. The t(1;22)(p13;q13), a translocation resulting in the OTT-MAL gene fusion and highly correlated with acute megakaryoblastic leukemia, has been to date detected exclusively in children, 96% of whom were younger than 24 months.36;37 Interestingly, almost all infants younger than 6 months had t(1;22) as a sole aberration whereas it was predominantly part of a complex karyotype in the majority of older children (p ¼ 0:00004).36 Another aberration hitherto detected only in infants aged 20 months or younger is t(7;12) (q36;p13). This subtle translocation, involving the ETV6 gene and almost always occurring together with trisomy 19, was often misdiagnosed as del(12p) in the past and only recently has been shown using fluorescence in situ hybridization (FISH) to be a consistent chromosome aberration in infant AML.38;39 In contrast, t(15;17)(q22;q12-21) and t(8;21)(q22;q22), the two most common reciprocal translocations in both adults and older children with AML (Table 1), have never been detected in infants aged less than 12 months.31 However, while the incidence of t(15;17) [and inv(16)(p13q22)/t(16;16)(p13;q22)] is similar in adults and older children, t(8;21) is twice as common in pediatric as in adult AML (Table 1). On the other hand, )5, del(5q) and other unbalanced structural abnormalities resulting in loss of material from 5q are much more frequent in adult than childhood AML. Likewise, both inv(3)(q21q26) and t(3;3)(q21;q26), which are found in 2% of adults, are extremely rare in children and have so far never been detected in a patient with de novo AML younger than 12 years.31
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Table 1 Frequencies of the more common cytogenetic abnormalities in adult and childhood AML Cytogenetic abnormality
None (normal karyotype) +8 )7/7q) )7 del(7q) Loss of (7q)f t(15;17)(q22;q21) )5/5q) )5 del(5q) Loss of (5q)f t(8;21)(q22;q22) inv(16)(p13q22)/ t(16;16)(p13;q22) )Y t/inv(11q23) t(9;11)(p22;q23) abn(12p) +21 abn(17p) del(9q) inv(3)(q21q26)/ t(3;3)(q21;q26) del(11q) t(9;22)(q34;q11) t(6;9)(p23;q34) Complex karyotype with P3 abn Complex karyotype with P5 abn
Cooperative Group Study (No. of patients) CALGBc ðn ¼ 1311Þ No. (%)
MRCd ðn ¼ 2337Þ No. (%)
SWOG/ECOGe ðn ¼ 609Þ No. (%)
Adults totala ðn ¼ 4257Þ No. (%)
Childrena;b ðn ¼ 1184Þ No. (%)
582 (44)
1096 (47)
244 (40)
1922 (45.1)
283 (23.9)
123 (9) 95 (7) 47 (4) 19 (1) 29 (2) 88 (7) 86 (7) 26 (2) 42 (3) 18 (1) 81 (6) 96 (7)
211 (9) 209 (9) 136 (6) 73 (3) NA 210 (9) 183 (8) 79 (3) 104 (4) NA 104 (4) 53 (2)
53 (9) 52 (9) NA NA NA 27 (4) 36 (6) NA NA NA 50 (8) 53 (9)
387 (9.1) 356 (8.4) 183 (5.0) 92 (2.5) 29 (2.2) 325 (7.6) 305 (7.2) 105 (2.9) 146 (4.0) 18 (1.4) 235 (5.5) 202 (4.7)
112 (9.5) 62 (5.2) 33 (2.8) 18 (1.5) 11 (1.3) 117 (9.9) 14 (1.2) 4 (0.3) 9 (0.8) 1 (0.1) 137 (11.6) 70 (5.9)
58 54 27 33 28 30 33 12
(4) (4) (2) (3) (2) (3) (3) (1)
NA 45 (2) NA NA 51 (2) NA 37 (2) 61 (3)i
20 (3) 42 (7)g NA NA NA 12 (2)h 17 (3) 12 (2)
78 (4.1) 141 (3.3) 27 (2.1) 33 (2.5) 79 (2.2) 42 (2.2) 87 (2.1) 85 (2.0)
28 (3.8) 155 (13.1) 54 (6.4) NA 60 (5.1) 5 (2.0) 33 (2.8) 0
12 (1) 10 (1) 8 (1) 135 (10)
NA 16 (1) 10 (<1) NA
NA 8 (1) 11 (2) 71 (12)
12 (0.9) 34 (0.8) 29 (0.7) 206 (10.7)
11 (1.3) 2 (0.2) 12 (1.0) 36 (14.3)
99 (8)
222 (9)
53 (9)
374 (8.8)
31 (5.2)
Abbreviations: CALGB, Cancer and Leukemia Group B; MRC, United Kingdom Medical Research Council; SWOG/ ECOG Southwest Oncology Group/Eastern Cooperative Oncology Group; abn, abnormality; NA, not available. a Percentages for particular abnormalities calculated using only those studies that provided relevant data. b Data from Leverger et al.166 (130 children aged from 2 months to 16 years diagnosed with untreated AML); Raimondi et al.167 (121 children diagnosed with de novo AML); Martinez-Climent et al.168 (115 children and adolescents aged from 0 months to 19.2 years diagnosed with de novo AML); Raimondi et al.169 (478 children and adolescents younger than 21 years diagnosed with de novo AML). c Data from Byrd et al.;20 the study comprised patients aged 15–86 years diagnosed with de novo AML. d Data from Grimwade et al.16 and Grimwade et al.;19 Grimwade et al.16 comprised 1272 patients aged 15–55 years, the majority of whom were diagnosed with de novo AML; up to 9.4% of patients had AML secondary to chemotherapy or radiotherapy or to an antecedent hematologic disorder. Grimwade et al.19 comprised 1065 patients aged 44–91 years, 817 of whom were diagnosed with de novo AML and 248 with AML secondary to chemotherapy or radiotherapy or to an antecedent hematologic disorder. e Data from Slovak et al.;18 the study comprised patients aged 16–55 years diagnosed with untreated AML. f Loss of 7q and Loss of 5q refer to unbalanced structural abnormalities, other than del(7q) and del(5q), that result in loss of material from the, respectively, 7q and 5q chromosome arms (e.g., unbalanced translocations, additions, isochromosome of 7p or 5p, etc.). g Category defined as “abn 11q”, might also include patients with other abnormalities of 11q including del(11q). h Including three cases with i(17)(q10). i Category defined as “abn 3q”, might also include patients with other abnormalities of 3q.
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Prognostic significance of cytogenetics in AML The Fourth International Workshop on Chromosomes in Leukemia (4IWCL) was the first large, prospective, multi-center study that established pretreatment karyotype as an independent prognostic factor in AML.8 Significant differences in complete remission (CR) rate, CR duration (CRD) and overall survival (OS) were demonstrated when the 716 patients were prioritized first according to the presence of t(8;21); then t(15;17); followed by )5 or del(5q), )7 or del(7q); concurrent occurrence of )5 or del(5q) and )7 or del(7q), and then abnormalities of 11q, +8 and +21. Patients without any of the above-mentioned abnormalities were classified according to the chromosome number [hypodiploid, pseudodiploid, diploid (normal) and hyperdiploid]. Cytogenetic findings so classified were independent prognostic factors for both duration of first CR and overall survival in the subset of 305 patients treated adequately.8 In the followup studies, a group of patients with abnormalities of 16q22 was added, all cases with aberrations of chromosomes 5 and 7 were combined into one category, and patients with +8 and +21 included in the hyperdiploid group.12;14;40 The multivariate analyses performed at the third follow-up of the 4IWCL, which then comprised 628 cases with de novo AML with a median follow-up of 14.7 years for patients alive, confirmed karyotype as an independent predictor of survival for all patients and for those 291 patients who received induction therapy that would be deemed standard by present-day criteria.14 Many studies, both smaller, performed in a single institution and large, collaborative multi-institutional ones, have confirmed that pretreatment karyotype constitutes an independent prognostic determinant for attainment of CR, CRD, risk of relapse and survival.9–11;13;15;16;18;20 Recently, three large collaborative studies proposed prioritization schemata that assign AML patients to one of the three risk groups, favorable, intermediate or adverse, based on pretreatment cytogenetic findings.16;18;20 The three cytogenetic risk systems share many common features, but differ with regard to some aspects (Table 2). In the MRC schema, any abnormality that is not classified as favorable or adverse, and is not accompanied by any additional chromosome aberration classified as favorable or adverse, is categorized in the intermediate-risk group. In contrast, both the Southwest Oncology Group/Eastern Cooperative Oncology Group (SWOG/ECOG) and Cancer and Leukemia Group B (CALGB) schemata explicitly
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categorize particular abnormalities into risk groups, and leave as not classified aberrations too infrequent to be analyzed. Moreover, MRC and SWOG/ECOG classify patients with a given abnormality into one of the three risk groups once, whereas CALGB provides risk-group assignment separately for probability of induction success, cumulative incidence of relapse (CIR) and OS. Consequently, in the CALGB schema, the same abnormality [e.g., t(6;9)(p23;q34)] may be categorized in the intermediate-risk group with regard to probability of achieving a CR, in the adverse-risk group with respect to OS and not being classified with regard to CIR, because too few patients attained a CR to be analyzed. Despite these differences, the inv(16)/t(16;16) and t(8;21), cytogenetic hallmarks of core-binding factor (CBF) AML, have been categorized in the favorable group by all three cytogenetic risk systems. However, SWOG/ECOG classified as favorable also patients with del(16q) whereas MRC and CALGB did not. We believe that patients with true del(16q) [i.e., deletions that do not represent misinterpreted inv(16) or t(16;16) and are usually found in AML with morphology other than that of acute myelomonocytic leukemia with abnormal eosinophils (AMML Eo)] should not be included in the favorable risk group, because del(16q) differs from inv(16)/t(16;16) at the molecular level, and has not been associated with a favorable outcome comparable to that of inv(16)/t(16;16).28;41;42 Consequently, the RT-PCR and/or FISH assays detecting CBFb-MYH11 gene fusion should be performed in all patients with del(16)(q22) to ensure that they do not harbor a misidentified inv(16)/ t(16;16).28 Moreover, while MRC and CALGB included all patients with t(8;21) in the favorable group, irrespective of whether t(8;21) was the sole aberration or occurred together with one or more secondary abnormalities, SWOG/ECOG classified as favorable only those t(8;21)-positive patients who did not have a complex karyotype with three or more abnormalities or a secondary del(9q). The latter abnormality has been reported as a poor risk indicator in patients with t(8;21) in one study,43 but this result was not corroborated by the MRC,16 CALGB,20 nor by other studies.8;10;44 Indeed, it has been consistently reported that for patients with t(8;21) and inv(16)/t(16;16), neither the presence of secondary abnormalities, including the most frequent in t(8;21)-positive patients )Y, )X and +8, nor a complex karyotype with three or more abnormalities adversely affects clinical outcome.8;10;16;20;44 Instead, adverse prognostic significance among t(8;21)-positive patients has been attributed to such factors as high initial white
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Table 2 Risk-group assignments of patients with selected chromosome aberrations in the 3 major collaborative cytogenetic studies of adult AML
Abbreviations: CALGB, Cancer and Leukemia Group B; MRC, United Kingdom Medical Research Council; SWOG/ ECOG Southwest Oncology Group/Eastern Cooperative Oncology Group; CR, complete remission; CIR, cumulative incidence of relapse; OS, 5-year overall survival; abn, abnormality; NA, not available. a For the convenience of the reader, risk group assignments of particular abnormalities are depicted in color: green for “favorable”, blue for “intermediate” and probably intermediate by virtue of being “other structural” or “other numerical” in the MRC classification, red for “adverse” and brown for “not classified” or “unknown”.
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Table 2 (continued) b Data from Byrd et al.;20 1213 adults with de novo AML analyzed. Patients with t(15;17) and t(9;22) were excluded from the analysis because therapy with ATRA or imatinib mesylate, respectively, is now considered a standard treatment option for these AML subtypes. All patients with t(8;21) and inv(16)/t(16;16) were classified in the favorable and those with t(9;11) in the intermediate risk categories regardless of whether t(8;21), inv(16)/t(16;16) and t(9;11) occurred alone or together with any secondary aberration or as part of a complex karyotype (defined as the presence of three or more aberrations). All other patients with a complex karyotype were classified as having adverse risk. Consequently, risk-group assignments of all other aberrations concerned only those patients who had a given aberration occurring alone or with only one additional abnormality other than t(8;21), inv(16)/t(16;16) and t(9;11).20 c Data from Grimwade et al.16 Of the 1612 patients analyzed, 1493 were diagnosed with de novo AML and 119 with secondary AML. The patient population included 340 children aged 0–14 years and 1272 adults aged 15–55 years. Patients with t(8;21), inv(16)/t(16;16) and t(15;17) were classified in the favorable risk group irrespective of whether t(8;21), inv(16)/t(16;16) and t(15;17) occurred alone or in conjunction with other abnormalities. Adverse abnormalities were classified as such regardless of whether they occurred alone or in conjunction with intermediate-risk or other adverse-risk abnormalities. Intermediate group comprised cytogenetic abnormalities not classified as favorable or adverse in the absence of accompanying favorable or adverse cytogenetic changes16 . d Data from Slovak et al.;18 609 adults aged 16–55 years with untreated AML analyzed. e Provided t(8;21) is not part of a complex karyotype or is not accompanied by del(9q). Both complex karyotype and del(9q) were classified in the adverse prognostic category. f Favorable for a group of 13 patients with del(9q) that included six who underwent SCT off-protocol; intermediate for non-transplanted patients treated with chemotherapy only. g Would be included in “abnormal 11q23” category. h Would be included in “abn 20q” category. i del(12p) classified as intermediate, other rearrangements affecting 12p presumably classified as unknown. j The abnormality accompanying +8 is other than t(8;21), t(9;11) and inv(16) or t(16;16). k Intermediate provided the abnormality accompanying +8 is not classified as favorable or adverse. l Would be included in “abn 3q” category. m Would be included in “abn 11q” category.
blood cell count (WBC) or absolute granulocyte count, the presence of granulocytic sarcomas, expression of the neural cell adhesion molecule CD56 on leukemic blasts and high WBC index.44 The WBC index [i.e., WBC (% of marrow blasts/100)], which divides patients into three subgroups (low index, <2.5; intermediate index, 2.5–20; high index, P 20) that differ with respect to clinical outcome, has been found in a recent large study to be the only prognostic factor for CRD, disease-free survival (DFS) and overall survival of patients with t(8;21) in multivariate analysis.44 The outcome of adults with CBF AML can be improved substantially by intensive post-remission therapy with high-doses of cytarabine (HDAC).17 Furthermore, patients with t(8;21) who received three or four cycles of HDAC on CALGB treatment protocols had superior projected 5-year DFS (71% versus 37%) and survival (76% versus 44%) rates compared with those who were administered only one cycle of HDAC followed by sequential treatments with cyclophosphamide/etoposide and mitoxantrone/diaziquone with or without filgrastim.45 The efficacy of repeated courses of HDAC for AML patients with t(8;21) has been confirmed by others.46 Likewise, the 5-year CIR was signifi-
cantly decreased in patients with inv(16)/t(16;16) receiving 3–4 cycles of HDAC as compared with those receiving one HDAC course (43% versus 70%) [Byrd et al., unpublished results]. This improved outcome is likely related to an increased sensitivity of the leukemic cells carrying inv(16)/t(16;16) or t(8;21) to cytarabine. Tosi et al.47 have shown a significant increase in incorporation of cytarabine into nuclear DNA in vitro and cytarabine induced apoptosis in blasts from patients with inv(16) compared to cells from patients with other chromosome aberrations or a normal karyotype. Another study revealed that samples from patients with inv(16)/t(16;16) and those with t(8;21) had the highest in vitro proliferative activity which was closely correlated with an increased incorporation of the active moiety of cytarabine, AraC triphosphate, into the DNA.48 The three major cytogenetic risk classifications agree that prognosis of patients with inv(3) (q21q26) or t(3;3)(q21;q26), )7 and a complex karyotype is poor (Table 2). The definition of complex karyotype differs, though, with the MRC characterizing this category as “the presence of a clone with at least five unrelated cytogenetic abnormalities”, and SWOG/ECOG and CALGB, as well
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as the German AML Study Group,49 as three or more abnormalities. A multi-center Italian study defined complex karyotype as “the presence of a clone with more than three cytogenetic abnormalities”.50 Byrd et al.20 compared the outcome of patients with three or four abnormalities [other than t(8;21), inv(16)/t(16;16) or t(9;11)(p22;q23)] with that of patients with five or more abnormalities. Although patients in the former group were younger and had significantly better CIR and 5-year OS than patients with five or more abnormalities, the CR rate and OS of patients with three or four abnormalities were significantly lower and the CIR significantly higher than those of patients included in the cytogenetically normal group. Only one patient in the group with three or four abnormalities remained in remission at 5 years. Thus, the authors concluded that these data justify combining patients with three or four abnormalities with patients who have five or more abnormalities into one complex karyotype category defined by the presence of three or more abnormalities.20 It has been striking in the CALGB series that all or almost all patients with )5, del(7q), )17/17p), )18, and )20 had a complex karyotype, thus precluding assessment of the prognostic significance of these abnormalities independently from complex karyotype. In the MRC study, the outcome of patients with del(7q) that was not part of a complex karyotype with P 5 abnormalities and was not accompanied by )5/del(5q) or abn(3q) did not differ significantly from outcome of patients with a normal karyotype.16 Their findings are consistent with earlier observations suggesting that patients with del(7q) without coexisting aberrations of chromosome 5 may have prolonged survival.40;51 In the CALGB series, most patients with del(5q) had a complex karyotype and very poor prognosis. However, a relatively small group of patients with del(5q) in a non-complex karyotype was classified as having intermediate risk with regard to CR rate and OS.20 Patients with various balanced abnormalities involving band 11q23 have been grouped into one cytogenetic category in both the SWOG/ECOG and MRC study, and classified, respectively, as having adverse and intermediate prognosis (Table 2). However, mounting evidence suggests that outcome of patients with translocations involving band 11q23 depends on the partner chromosome involved, with t(9;11)(p22;q23)-positive patients having a more favorable prognosis that places them in the intermediate-risk group.20;52 Remarkably, a recent study of 298 infants and children with de novo AML found that t(9;11) was the most important, favorable prognostic factor for patients
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treated on protocols used at St. Jude Children’s Research Hospital.53 Zwaan et al.54 suggested that the superior outcome of the t(9;11)-positive patients may be explained by enhanced sensitivity of their leukemic blasts to several chemotherapeutic drugs. In this study, bone marrow or blood samples from children with AML and t(9;11) were significantly more sensitive in vitro to cytarabine, etoposide, the anthracyclines and 2-chlorodeoxyadenosine than samples from patients with various other chromosome aberrations. When compared with a subgroup of childhood AML patients harboring other 11q23 translocations, including t(6;11)(q27;q23), t(9;11)-positive samples were significantly more sensitive for cytarabine and doxorubicin, and borderline more sensitive for etoposide.54 The survival of adults with t(6;11) and t(11;19)(q23;p13.1) studied by CALGB was significantly shorter than that of the cytogenetically normal group, and, consequently, t(6;11) and t(11;19)(q23;p13.1) were assigned to the adverserisk group for OS. For patients with other, less frequent 11q23 translocations, the definitive assignment of risk category will be possible only once enough patients are analyzed in large prospective studies.20 The most common trisomies in de novo AML are, in decreasing order of frequency, +8, +22, +13, +21 and +11.20 Trisomy 22 is a non-random secondary aberration accompanying inv(16)/t(16;16) and is rarely seen as the only chromosome abnormality.31 Although each of the remaining trisomies can be found as a secondary aberration, +8, +13, +11 and +21 are also detected recurrently as the only (isolated) karyotypic changes at diagnosis, with a frequency among adults with de novo AML of 4% for sole +8, 1% each for sole +13 and +11, and 0.4% for sole +21.55 With regard to the impact of recurrent trisomies on clinical outcome, most data have been gathered for trisomy 8, but results have been somewhat inconsistent. CR rates of patients with +8 have differed widely, from 29%11 to 91%,51 as have CRD and survival among studies,32 and consequently patients with +8 have been classified either in the intermediate or adverse-risk category. In some reports, this cytogenetic group included both patients with isolated +8 and those who in addition to +8 had other aberrations that may have affected response to treatment and outcome. It has been repeatedly shown that prognosis of AML patients with +8 indeed depends on whether +8 occurs as an isolated abnormality or is accompanying other aberrations.16;20;56–58 In the latter situation, +8 does not appear to adversely affect the favorable outcome of patients with t(15;17), inv(16)/t(16;16) and t(8;21).16;20;56;57 In contrast,
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patients with +8 and a complex karyotype and/or such unfavorable aberrations as del(5q) or )7 usually have very poor outcome. In one recent study, patients with +8 and unfavorable chromosome aberrations (according to the SWOG/ECOG classification) had significantly worse OS than other patients with unfavorable aberrations who lacked +8 in their karyotype.58 On the other hand, isolated +8 has been considered to bestow either intermediate or unfavorable prognosis (Table 2). The conflicting results might be associated with variation in the age of the patients analyzed in different series,30 but may also stem from differences in proportion of patients who underwent stem cell transplantation (SCT). Farag et al.55 reported that prognosis of patients with an isolated +8, +11, +13 and +21 was dependent on SCT. While these isolated trisomies constituted an independent adverse prognostic factor for OS in non-transplanted patients, in those receiving SCT in first remission, the 5-year OS was not different than that of transplanted patients with a normal karyotype thus changing the risk stratification of isolated trisomies from adverse to intermediate.55 Prospective studies are required to confirm this finding. Patients with a normal karyotype are the single largest cytogenetic subset of adult AML. They are classified in the intermediate prognostic category, because their CR rates, CRD and survival probabilities are usually lower than those of adequately treated patients with t(8;21), inv(16) or t(15;17), but higher than patients with unfavorable chromosome aberrations.16;20;51 However, this group is highly heterogeneous at the molecular level, and is likely composed of subsets with varying prognoses. A small fraction of patients determined to have a normal karyotype by conventional cytogenetic analysis may carry gene fusions typically associated with recurrent translocations or inversions. Occasionally, this happens because the presence of a chromosome aberration is not recognized by the cytogenetic laboratory, especially when the quality of chromosome preparation is suboptimal and the missed rearrangement subtle, e.g., t(11;19)(q23;p13.1), t(15;17), or inv(16). However, in other patients, the gene fusion results from a cryptic rearrangement involving segments smaller than the length of a single band and thus unrecognizable by standard cytogenetic analysis. For instance, cryptic insertions of a very small segment from 17q containing the RARa gene into the locus of the PML gene on chromosome 15q have been repeatedly detected using FISH in patients with acute promyelocytic leukemia (APL) and a normal karyotype.59 Grimwade et al.59 estimates that insertions leading to formation of the PML-RARa fu-
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sion gene, most of which are cryptic, account for approximately 4% of all patients with APL. Because marrow of patients with cryptic PML-RARa fusions has a typical APL morphology, and such patients share the favorable prognosis of those with the standard t(15;17),59 karyotypically normal patients suspected of having APL should be tested for the presence of the PML-RARa fusion gene by reverse transcription-polymerase chain reaction (RT-PCR), Southern blot analysis or/and FISH, and, if found positive, treated with regimens containing alltrans-retinoic acid (ATRA). Similarly, rare patients with the CBFb-MYH11 fusion gene but without microscopically detectable abnormalities of chromosome 1660;61 and those with the RUNX1-CBFA2T1 (AML1-ETO) fusion gene and normal chromosomes 8 and 2162;63 should likely be grouped together with other patients with CBF AML. Of note, as demonstrated by two large prospective series,28;63 patients truly positive for the CBFb-MYH 11 and Runx1-CBFA2T1 gene fusion transcripts in the absence of cytogenetically detectable aberrations of chromosomes 16, and 8 and 21, respectively, are rare and their frequencies are much lower than those indicated by two retrospective studies.64;65 . It is also rather unlikely that a large proportion of cytogenetically normal patients with de novo AML harbor hidden aberrations detectable only by FISH-based techniques. In a series of 102 adult AML patients tested by FISH using a comprehensive set of genomic DNA probes capable of detecting major AML-associated aberrations, only three patients were found to harbor, respectively, inv(16), )Y and del(12p).66 In retrospect, only the del(12p) appeared to represent a truly cryptic aberration.66 Cuneo et al.67 did not find any occult chromosome anomaly among 55 adults with de novo AML aged 15–60 years who were studied by FISH using probes detecting )5, )7, +8, and deletions of 5q31, 7q31, 12p13/ETV6, 17p13/TP53 and 20q11. The authors did identify, however, chromosome aberrations in interphase cells of 3 of 21 (14%) AML patients aged 62–80 years and in 5 of 6 (83%) patients with AML secondary to an antecedent MDS. In four of these patients, FISH analysis of metaphase cells revealed submicroscopic del(5)(q31) (three cases) and del(7)(q31) (one case).67 To date, application of multi-color FISH-based techniques that use chromosome painting probes, such as spectral karyotyping (SKY) and multiplex FISH (M-FISH), to study AML patients with a normal karyotype has failed to discover any novel hidden rearrangement, akin to the cryptic t(12;21)(p13;q22) in childhood pre-B ALL, which, due to the juxtaposition of similarly banded regions, can be discerned microscopically only by FISH.68–71 On the other hand, a new recur-
Rearrangementa
Age group
Prognostic relevanceb
No. pts with/no. pts without rearrangement
Reference number
11/87 7/61 15/30
72
Normal karyotype PTD MLL
Adults
PTD MLL
Adults
PTD MLL
Adults 16–60 years
PTD MLL FLT3 ITD
Adults 16–60 years Adults 16–60 years
FLT3 Asp835
Adults 16–60 years
FLT3 ITD versus FLT3 Asp835 FLT3 ITD
Adults 16–60 years
FLT3 ITD
Adults 20–59 years
FLT3ITD=
Adults 20–59 years
FLT3ITD=WT
Adults 20–59 years
Adults
FLT3ITD= versus FLT3ITD=WT Adults 20–59 years
CR rate and OS: no significant difference CRD: significantly shorter for PTD MLL+ patients Median survival: significantly shorter for PTD MLL+ patients compared with that of age-matched karyotypically normal control group Relapse-free interval: significantly shorter for PTD MLL+ patients compared with that of age-matched karyotypically normal control group CR rate and OS: no significant difference CRD: significantly shorter for PTD MLL+ patients EFS: significantly shorter for PTD MLL+ patients CR rate: no significant difference CRD: significantly shorter for FLT3 ITD+ patients OS: significantly shorter for FLT3 ITD+ patients CR rate and OS: no significant difference CRD: no significant difference CR rate and OS: no significant difference CRD: no significant difference CR rate: no significant difference OS: significantly shorter for FLT3 ITD+ patients CRD: significantly shorter for FLT3 ITD+ patients CR rate and OS: no significant difference DFS: significantly shorter for FLT3 ITD+ patients CR rate: no significant difference OS: significantly shorter for FLT3ITD= patients DFS: significantly shorter for FLT3ITD= patients CR rate and OS: no significant difference DFS: no significant difference CR rate: no significant difference OS: significantly shorter for FLT3ITD= patients DFS: no significant difference
73
Cytogenetics in acute leukemia
Table 3 Molecular genetic rearrangements reported to affect clinical outcome of AML patients with a normal karyotype or those classified in the intermediate prognostic category
9/17
18/203 16/158 5/25 67/125 47/95 67/125 28/125 23/95 67/28 47/23 16/37 16/37 8/28 23/59 17/51 8/59 8/59 6/51 15/59 11/51 8/15 8/15 6/11
75 29 84
84 84 88
80 80
80 80
123
124
Table 3 (continued) Rearrangementa
Prognostic relevanceb
Age group
No. pts with/no. pts without rearrangement
Reference number
66/217 46/139 46/139 76/172 29/192
85
Intermediate-risk prognostic groupc FLT3 ITD
Adults <60 year
FLT3 ITD or FLT3 TKD Adults <60 year FLT3 ITD mutant/wt allele Adults <60 year ratio >0.78
FLT3 ITD mutant/wt allele Adults <60 year ratio >0 6 0.78 FLT3 ITD mutant/wt allele Adults <60 year ratio >0.78 versus >0 6 0.78
Adults
FLT3 ITD FLT3 ITD
Adults 15–74 years Adults
FLT3 ITD or FLT3 Asp835
Adults
FLT3 ITD FLT3 ITD N-RAS point mutation GSTM1 or/and GSTT1
Adults Adults Adults Adults
16–88 15–85 15–85 19–75
years years years years
85 85
23/152 23/152 30/192 25/152 25/152 29/30
85
85
23/25 23/25 118/288 109/251 44/94 10/34 15/65 17/54 27/65 26/54 15/40 123d 123d 51/44 31/36 51/44
86
77 89 89 78 76 76 94
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FLT3 ITD
CR rate: no significant difference DFS: no significant difference Probability of relapse: significantly higher for FLT3 ITD+ patients OS: significantly shorter for FLT3 ITD+/FLT3 TKD+ patients OS: significantly shorter for FLT3 ITD patients with mutant/ wt allele ratio >0.78 DFS: significantly shorter for FLT3 ITD patients with mutant/wt allele ratio >0.78 Probability of relapse: significantly higher for patients with mutant/wt allele ratio >0.78 OS: no significant difference DFS: no significant difference Probability of relapse: no significant difference OS: significantly shorter for FLT3 ITD patients with mutant/wt allele ratio >0.78 DFS: significantly shorter for FLT3 ITD patients with mutant/wt allele ratio >0.78 Probability of relapse: significantly higher for patients with mutant/wt allele ratio >0.78 EFS: significantly shorter for FLT3 ITD+ patients OS: no significant difference DFS: no significant difference OS: significantly shorter for FLT3 ITD+ patients EFS: no significant difference DFS: no significant difference EFS: no significant difference DFS: no significant difference DFS: significantly shorter for FLT3 ITD+ patients OS: significantly shorter for FLT3 ITD+ patients OS: significantly shorter for patients with N-RAS point mutation CR rate: significantly lower for GSTM1 /GSTT1 patients EFS: significantly shorter for GSTM1 /GSTT1 patients OS: significantly shorter for GSTM1 /GSTT1 patients
a
Unless otherwise indicated, all comparisons are between patients with a given gene rearrangement and those without (wild-type allele); PTD MLL, partial tandem duplication of the MLL gene; FLT3 ITD, internal tandem duplication of the FLT3 gene; FLT3ITD=WT cases of AML with the FLT3 ITD in combination with the FLT3 wildtype (WT) allele; FLT3ITD= , cases of AML with the FLT3 ITD lacking a FLT3 WT allele; FLT3 Asp835, activating point mutations of D835 within the activation loop of the FLT3 gene; FLT3 TKD, activating point mutations in codon 835 or 836 of the FLT3 gene; GSTM1 or/and GSTT1 , homozygous deletions resulting in null genotypes at the glutathione S-transferase GSTM1 or/and GSTT1 loci. b CR, complete remission; CRD, CR duration; EFS, event-free survival; OS, overall survival. c Intermediate-risk prognostic group defined as follows: Thiede et al.85 – normal karyotype and chromosome aberrations other than those conferring high risk [i.e., )5/del(5q),)7/del(7q), hypodiploid karyotypes (besides isolated )Y or )X), inv(3q), abn 12p, abn 11q, +11, +13, +21, +22, t(6;9); t(9;22); t(9;11); t(3;3) and multiple aberrations ( ¼ 3 independent aberrations)] and those conferring low risk [i.e., t(8;21) and t(8;21) combined with other aberrations]; Schnittger et al.86 – normal karyotype and aberrations other than t(15;17), t(8;21), inv(16)/t(16;16), t(11q23), t(6;9), t(1;3), inv(3)/t(3;3), t(3;12), t(3;21), t(8;16) and )5/)7/7q); Abu-Duhier et al.77 and Voso et al.94 – normal karyotype and +8, +21, +22, del(7q), del(9q), abnormal 11q23 and all other structural/numerical abnormalities without additional favorable [i.e., t(8;21), t(15;17) and inv(16)] or adverse [i.e., )5, )7, del(5q), abnormal 3q and complex karyotype with 5 or more abnormalities] cytogenetic changes; Moreno et al.89 – normal karyotype and aberrations other than )5/del(5q), )7/del(7q), abn 3q, complex aberrations ( P 3 independent aberrations), t(9;22), t(6;9); t(8;21) and inv(16); Rombouts et al.78 – normal karyotype and aberrations other than t(15;17), t(8;21), inv(16), t(6;9), alterations in the 11q23 region, del(5), del(7) or deletions of the q-arms of these chromosomes, and complex karyotype with more than 3 abnormalities; Kioyi et al.76 – normal karyotype and aberrations other than t(8;21), inv(16), t(9;22), 11q23 alterations, del(5) or del(7). d Intermediate-risk prognostic group included 123 patients, but numbers of patients with and without FLT3 ITD and N-RAS point mutation were not provided.
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rent AML-associated cryptic translocation, t(5;11)(q35;p15.5), was recently discovered by Brown et al.71 who used a 12-color FISH assay (termed M-TEL) capable of detecting subtle subtelomeric rearrangements that are below the resolution of G-banding. The t(5;11), which results in the NUP98-NSD1 gene fusion, was found in 2 of the 61 (3%) patients with a normal karyotype.71 The presence of an exclusively normal karyotype in a diagnostic marrow sample does not mean that cytogenetically normal leukemic blasts harbor no acquired genetic alterations. A variety of genetic abnormalities discernible only by molecular genetic techniques, such as Southern analysis, RTPCR or direct sequencing, have been reported. These include partial tandem duplication of the MLL gene (PTD MLL);29;72–75 internal tandem duplication of the FLT3 gene (FLT3 ITD);76–89 activating point mutations of D835 within the activation loop of the FLT3 gene (FLT3 Asp835 mutation);84;85;89;90 point mutations of the NRAS,76;81 CEBPA,91 and RUNX1 (AML1) genes;92 deletions and point mutations of the PU.1 gene;93 homozygous deletions resulting in null genotypes at the glutathione S-transferase GSTM1 and GSTT1 loci;94 and overexpression of the BAALC,95 and EVI1 genes.96 The molecular abnormalities shown to influence prognosis of AML patients with a normal karyotype and of those comprising the intermediate-risk cytogenetics group (which usually contains a large proportion of karyotypically normal patients) are provided in Table 3. All four studies examining the prognostic significance of the PTD MLL in patients with a normal karyotype agree that the presence of PTD MLL significantly shortens the patients’ CRD or event-free survival (EFS);72–75 in one study, OS of patients with PTD MLL was also significantly shorter than that of an age-matched karyotypically normal control group.73 The prognostic impact of FLT3 ITD, the most common gene rearrangement in AML, is more controversial. Some studies investigating patients with a normal karyotype or those comprising the intermediaterisk cytogenetics group found no significant difference in the DFS,85;86;89 EFS89 and OS80;86 between patients with and without FLT3 ITD. In other studies, patients with FLT3 ITD had significantly worse CRD,84;88 DFS,78;80 EFS86 and OS.76;77;88 These differences among studies may be attributed to several factors, including varying numbers of patients studied, differences in age of the patients, differing definitions of the intermediate-risk group (see footnote to Table 3) and thus disparate proportions of patients with particular “intermediate-risk” chromosome aberrations, and distinct induction and post-remission therapies
126
administered to patients in different studies. Additionally, some studies did not separate patients who had FLT3 ITD in combination with the FLT3 wild-type allele from those with the FLT3 ITD who lacked a FLT3 wild-type allele. Two groups have recently demonstrated that only patients with the latter genotype (or a high mutant/wt ratio, greater than 0.78) had a significantly shorter DFS and OS than patients without FLT3 ITD.80;85 Importantly, FLT3 ITD is also common in APL patients with t(15;17), but hitherto has not been shown to predict prognosis in these patients.87;88;97 This underscores the importance of assessing prognostic value of novel gene rearrangements in the context of cytogenetically defined groups of patients with AML.
Acute lymphoblastic leukemia The first comprehensive study of the cytogenetics of adult ALL was the Third International Workshop on Chromosomes in Leukemia,21 which showed that adult ALL cytogenetics have biologic and prognostic significance. Four cytogenetic classification schemata were defined: (1) according to the presence of normal and abnormal metaphases: NN – all metaphase cells normal, NN + Ncl – normal cells and non-clonal abnormalities, AN – normal cells and clonal abnormalities and AA – only abnormal metaphases; (2) according to the modal chromosome number, dividing the patients with an abnormal karyotype into six groups, 6 35, 36–45, 46 (pseudodiploid), 47–50, 51–60 and >60 chromosomes, and grouping patients with no clonal abnormalities in the “46 normal” category; (3) normal, abnormal with a translocation identified and abnormal without an identified translocation; (4) the Lund classification.21 The Lund classification was comprised of 10 hierarchical groups, t(9;22)(q34;q11.2) or a Philadelphia chromosome (Ph), t(4;11)(q21;q23), t(8;14)(q24;q32) or del(8q), other 14q+ abnormalities, del(6q), and normal karyotype, with the remaining cytogenetically abnormal patients classified by modal chromosome number as hypodiploid, pseudodiploid, low hyperdiploid (47–50 chromosomes) and high hyperdiploid (>50 chromosomes). By the Lund classification, survival was longest for patients with high hyperdiploidy and for those with normal karyotypes. Patients with t(4;11), t(8;14), 14q+, hypodiploidy and low hyperdiploidy had the shortest survival times. Remission rates were lowest in patients with hypodiploidy, t(9;22), t(4;11), t(8;14) and 14q+. Importantly, the karyotype was independent of other known prognostic factors in a
zek et al. K. Mro
multivariate analysis, although other clinical features were associated with modal number. Older age was more common in patients with low hyperdiploidy, and patients with a normal karyotype were more likely to be younger and have T-cell disease. Higher WBC was associated with pseudodiploidy, hypodiploidy, t(4;11) and t(9;22). With 6year follow-up, patients with FAB L1, WBC P50,000/ll and high or low hyperdiploidy, normal metaphases or with a del(6q) were most likely to have achieved a cure; however, the karyotype did not add prognostic significance for disease-free survival to age, WBC and immunophenotype.24 Additional reports have confirmed these observations, the clinical significance of some abnormalities has been further elucidated, and the molecular basis and biological consequences of many aberrations have been described. Below we will summarize the significance of the cytogenetics of adult ALL described since the International Workshops, particularly the more common abnormalities. Mature B-cell ALL will not be considered. The frequency of abnormal karyotypes in adult ALL is slightly higher than in pediatric ALL; 64–85%25;26;98–100 and 60–69%101;102 of successful cases, respectively. The frequencies of different ploidy groups and some aberrations, however, differ between children and adults, as does treatment outcome (Table 4). The overall survival rate for adults is 22–38%,25;99;103 whereas children have a 75–80% survival rate.104–107 The frequency of Ph+ ALL increases with age, whereas the frequency of high hyperdiploid ALL decreases with age. Less than 6% of children with ALL have a t(9;22), whereas up to 40% of adults aged P 40 years with ALL are Ph+, a poor prognostic feature regardless of age.25;108 In contrast, less than 12% of adults23;25;98;100 but 25% of children101;102;109;110 have high hyperdiploidy, a good prognostic feature. The different outcomes for adults compared with children with ALL may be partially a result of the different frequencies of specific abnormalities. A t(9;22) is the most common recurring abnormality in adult ALL, occurring in 11–29% of patients included in series comprising both B- and TALL,25;26;98 and in 37% of patients diagnosed with Bcell precursor ALL (c-ALL and pre-B ALL) who were studied by RT-PCR.103 Most (97%) Ph result from a standard t(9;22); only 3% are variant translocations. Patients with Ph+ ALL present with a high WBC (23,500/ll versus 11,550/ll),103 have B-lineage disease, a typical immunophenotype of CD34+/ CD10+/CD19+,111 and myeloid markers in up to 71% of adult cases.112 The remission rate usually is similar to the overall ALL remission rate,113 although has been lower in some studies.103 Progno-
Cytogenetics in acute leukemia
127
Table 4 Frequencies of cytogenetic aberrations in adult and childhood ALL and their prognostic relevance Abnormality
None (normal chromosomes) High hyperdiploidy Low hyperdiploidy Near-triploid/ near-tetraploid
Adults
Children
Frequencya
Prognosisa
Frequencya
Prognosisa
15–36%25;26;98–100;128
Intermediate– good26;98;127 Good25;26;98;100;127 Good25 Intermediate25
31–40%101;102
Intermediate– good101;102 Good101;102 Intermediate101;102
2–11%23;25;98;100 10–15%25;98;129 3–5%25;100 Near-triploidy: 3%98 Near-tetraploidy: 2%98
23–26%101;102;109;110 10–11%101;102 1%110;126
Near-triploidy: Poor98 Near-tetraploidy: Excellent98
Near-triploidy: Good102;126 Near-tetraploidy: Adverse126 Intermediate102 Intermediate101;102 Modal number ¼ 45: Intermediate101 Modal number ¼ 30–45: Intermediate102 Modal number <45: Poor101 Poor101;102 Poor102;171;172
Pseudodiploidy Hypodiploidy
31–50%25;98;100;129 4–9%25;98;100;129
Poor25 Poor25;98
18–26%101;102 6%101;102
Near-haploidy t(9;22)(q34;q11.2)
Not known Poor26;103;129;131;170
<1%101;102 2–6%102;108;171
t(4;11)(q21;q23) t(1;19)(q23;p13.3)
Rare100 11–29% overall25;26;98 37% of B-ALL by RT-PCR100 3–7%25;26;98;100;130;131 2–3%25;98;100;135
2%102 d 4–5%174;175
t(12;21)(p13;q22)
0–3%150–152
Poor26;133 Poor98;100;127;137 Good25 Not known
20–25%147–149
Abnormal 9p
6–30%25;26;98;100;129
Intermediate25;26;98
7–11%102;176
Abnormal 12p
4–6%25;26;98;100
7–9%102;177
del(6q)
3–16%25;26;129
6–9%102;178
Not prognostic102;178
del(7p)/del(7q)/ monosomy 7 del(5q) +8c 14q11
6–11%26;98b
Favorable25;26 Unfavorable98 Not prognostic132 Intermediate98 Adverse145 Not prognostic155 Poor26;132 Not prognostic155 Poor26 Excellent, especially t(10;14) (q24;q11)26;98
Poor173 Excellent102 Intermediate174 Good in most studies147–149 Adverse138;176 Not prognostic102 Not prognostic102;177
4%110
Adverse110
1%179 2%Unpublished data from CCG c 3–4% overall or 17–22% of T-ALL180;181
Adverse179 Unknown Not Prognostic180;181
t(10;14)(q24;q11)
<2%155 10–12%26;98 5–7% overall or 26% of T-ALL26;98 2–3%98
a
Superscript numbers denote reference numbers identifying articles that contain data on a given abnormality frequency and clinical outcome of patients with this aberration. b In Wetzler et al.26 , only patients with monosomy 7 were included. c Non-high hyperdiploid cases; CCG, Children’s Cancer Group. d Excluding infants.
sis is poor with traditional therapy and may be increasingly poor with increasing age.100 However, two studies showed a markedly reduced risk of relapse and increased survival for younger Ph+ ALL
patients who underwent allogeneic SCT from HLA-matched related donors in first CR and were conditioned with regimens that included fractionated total body irradiation and high-dose
128 etoposide.114;115 Additionally, newer biological therapy with imatinib mesylate, a drug powerfully effective in CML, another leukemia with t(9;22), also may be effective in ALL when combined with chemotherapy or other approaches. This drug competitively inhibits the abnormal protein produced in Ph+ disease,116 and may help to improve the outcome in Ph+ ALL. Early studies of Ph+ patients with ALL or CML with lymphoid blast crisis who had no response to standard treatment or who relapsed after standard treatment showed an initial 70% response rate to this therapy, but the responses have been of short duration.4 In a subsequent study of 48 patients with relapsed or refractory Ph+ ALL, imatinib induced complete hematologic and marrow responses in 29% of patients but development of resistance and subsequent disease progression were rapid, thus necessitating further studies of the efficacy of imatinib in combination with other chemotherapeutic agents in Ph+ ALL patients.116 The genes involved in the t(9;22) are ABL on chromosome 9 and BCR on chromosome 22. The 50 portion of BCR is joined to the 30 portion of ABL to form a chimeric gene, BCR-ABL. Two different breakpoints in BCR give rise to proteins of different sizes. The major breakpoint cluster region (M-bcr) spans BCR introns 12–16, and when fused with ABL results in a 210-kDa protein. This is the typical rearrangement in CML and in 24–50% of adult Ph+ ALL,113;117 but is rare in pediatric Ph+ ALL.118 The minor breakpoint (m-bcr) occurs 50 of the M-bcr in the first intron of BCR, and when fused with ABL, codes for a 190-kDa protein.119 This breakpoint occurs in 50–77% of adult Ph+ ALL103;117 and over 90% of pediatric Ph+ ALL.120 The proportion of patients with breakpoints in M-bcr increases with increasing age.121 Both proteins are tyrosine kinases, but the m-bcr product has stronger transforming potency in transfection assays and in transgenic mice.117 Comparisons of adults with Ph+ ALL with m-bcr and M-bcr breakpoints rarely show differences in the presence of additional cytogenetic aberrations, WBC, age, or outcome;117;122;123 although one study showed that patients with M-bcr breakpoints were older103 and another study of allogeneic transplants showed that patients with an m-bcr breakpoint had an increased risk of relapse compared with those with an M-bcr breakpoint.124 Cytogenetic abnormalities in addition to a t(9;22) occur in 41–86% of adult ALL at diagnosis;117;122 most frequently loss of a chromosome 7 or its short arm (7p), an additional der(22)t(9;22), abnormalities of 9p, gain of a chromosome 8 or X, duplication of 1q and high hyperdiploidy.117;122 No pretreatment clinical factors have been associated
zek et al. K. Mro with the presence of additional abnormalities.122 Outcome has been reported as no different,122;125 or dependent on the specific aberration in addition to t(9;22), with hyperdiploid patients having a better outcome and those with monosomy 7 or deletions of 9p having a particularly poor outcome.117 High hyperdiploidy is rare in adult ALL (2–11%)23;25;98;100 compared with pediatric ALL (25%).101;102;109;110 Near-triploidy and near-tetraploidy are more frequent in adult than in pediatric ALL.25;100;110;126 High hyperdiploidy in adults is associated with younger age and normal WBC. Outcome for adult high hyperdiploid patients is somewhat improved compared with other ploidy groups, but does not show the excellent outcome observed in children.25;26;98;100–102;109;110;127 Structural abnormalities are frequent in high hyperdiploid cases, most commonly a t(9;22). Patients with high hyperdiploidy and t(9;22) have outcomes similar to other Ph+ patients.98;125 Clinical features and outcome in patients with high hyperdiploidy and other recurring translocations also are similar to those of patients with the translocations and pseudodiploidy or low hyperdiploidy.25 Low hyperdiploidy is distinct from high hyperdiploidy both in chromosome pattern and in clinical features. Recent studies show an improved outcome for patients with low hyperdiploidy compared with patients with 6 46 chromosomes.25 Normal, diploid karyotypes occur in 15–36% of adult ALL25;26;98–100;128 and are associated with an intermediate26 or an improved outcome.26;98;126 Only 4–9% of adult ALL are hypodiploid.25;98;100;129 Patients with hypodiploidy are somewhat younger than those with normal karyotypes, and most have B-lineage ALL. Severe hypodiploidy with modal numbers of 30–44 is more frequent in adult ALL than is near-haploidy with chromosome numbers below 30. These patients have a poor outcome.25;98 Recurrent structural abnormalities occur in 60–70% of pseudodiploid patients,25;98 and the prognosis of these patients reflects the prognosis of the recurrent abnormality. When all pseudodiploid patients are considered, their outcome is worse than that of patients with normal chromosomes or with hyperdiploidy.25 A common recurring translocation in adult ALL is t(4;11)(q21;q23). Although particularly common in infant ALL, it also occurs in 2% of pediatric102 and 3–7% of adult ALL.25;26;98;100;130;131 It is associated with an immunophenotype of TdT+/HLA-DR+/ CD34+/CD10)/CD19+,111;132 a high WBC, a high frequency of circulating blasts and an increased frequency of central nervous system disease.132 The t(4;11) predicts a poor prognosis.26;133 Secondary abnormalities in addition to t(4;11) are
Cytogenetics in acute leukemia
common, particularly i(7)(q10) and trisomy 6; outcome does not appear to differ for these patients compared with those with t(4;11) as a sole abnormality.130 The gene involved on chromosome 11 is MLL, which has multiple translocation partners. In t(4;11), MLL is rearranged with AF4, the most common partner gene of the MLL gene in ALL. Other 11q23 (MLL) translocations also occur and overall, MLL translocations are found in up to 7% of adult ALL,26;98 occurring in both de novo and in therapy-related disease.134 Translocation (1;19)(q23;p13.3) occurs in 2–3% of adult ALL25;98;100;135 and, as in pediatric ALL, can be either balanced or an unbalanced der(19)t(1;19) with two normal chromosomes 1. Most cases are pseudodiploid, although some are high hyperdiploid. The rearrangement results in a chimeric E2APBX1 gene in 95% of cases with 6 50 chromosomes, but in only 25% of cases with >50 chromosomes.136 Nearly all cases are pre-B ALL; that is, the cells have cytoplasmic mu immunoglobulin, but not surface immunoglobulin, and are CD9+/CD10+/ CD19+/CD22+/CD34). Low WBC is common, and the patients tend to be younger.98 Outcome is poor in most studies of adults,98;100;127;137 although not all.25 Abnormalities of 9p that result in loss of genetic material [abn(9p)], including deletions, unbalanced translocations and chromosomal losses, are recurrent in ALL. Abn(9p) occurs in both T- and Blineage ALL, with some studies indicating a higher frequency in T-lineage138 and some in B-lineage ALL.139 Abn(9p) appears to be similar in adult and pediatric ALL.132 Cytogenetically, 6–30% of adult ALL patients have an abn(9p), and more show loss by molecular technologies.25;26;98;100;129 Prognosis is intermediate.25;26;98 Interestingly, several dicentric chromosomes recur in ALL; most involve chromosome arm 9p and result in partial loss of the p-arm. Dic(9;20)(p11–13;q11) has a good prognosis,140 as does dic(9;12)(p11–13;p11–12) even though most patients with the latter have at least one adverse feature.141 Dic(9;12) is more frequent in younger patients and is rare in adults.132 Abnormalities in addition to dic(9;12) are common, but complex karyotypes are rare.141 Isochromosome 9q, which results in loss of 9p, also is recurrent in ALL.142 The most common region of loss is 9p21, and deletion of the p16INK4a gene (also called CDKN2A or MTS1) appears to be the significant loss. Both hemizygous and homozygous deletions occur, with no apparent difference in clinical features or outcome.139 The neighboring gene p15INK4b (MTS2) also is lost frequently and may contribute to the pathogenesis of ALL.139 Loss of expression of p15INK4b and of p16INK4a may occur through methylation of the re-
129
maining allele in some patients with hemizygous loss.143;144 Recent studies have failed to show a prognostic significance for del(6q)132 , have shown an intermediate outcome,98 or an adverse outcome.145 Patients with del(6q) tend to be younger than those with normal 6q.145 Deletion of 6q may be a sole abnormality or part of a complex karyotype. An isolated del(6q) has been associated with hyperleukocytosis and T-ALL.145 Breakpoints vary, with band 6q21 most frequently cited as containing the smallest region of deletion.132;145 It is assumed that an as yet unidentified tumor suppressor gene is lost as a result of the deletion. Rearrangements of 12p occur in 4–6% of adult ALL,25;26;98;100 frequently as part of a complex karyotype. Translocations with many different partner chromosomes have been reported, as well as partial loss of 12p, resulting from both deletions and unbalanced translocations. Patients with 12p abnormalities tend to be younger and have a favorable prognosis in some studies,25;26 but not in all.98 One recently described aberration of 12p is the cryptic t(12;21)(p13;q22), resulting in fusion of ETV6 (TEL) and AML1 (RUNX1, CBFA2), in pre-B ALL.146 It occurs in up to 25% of children aged 1–10 years,147–149 but in only 0–3% of adult ALL.150–153 In children, it usually is associated with an excellent prognosis,147;149;151 although not always.154 Because it is infrequent in adult ALL, its prognostic significance in adults remains to be determined. Other less frequently recurring cytogenetic abnormalities in adult ALL include trisomy 8 and partial or complete monosomy for chromosomes 5 ()5/5q)) and 7 ()7/7q)).26;132;155 They are often secondary to t(9;22) and in those cases are associated with the poor outcome of Ph+ ALL.155 Patients with )5/5q) tend to be younger and frequently have T-ALL.156 Cases with )5/5q)/)7 without t(9;22) have remission and survival rates similar to patients with normal chromosomes 5 and 7,98;155 although in another study patients with )7 and +8 who were Ph-negative fared as poorly as the Ph+ patients.26 T-ALL occurs in 14–28% of adult ALL.25;26;98;100 Although the cytogenetics of T-lineage and B-lineage ALL overlap, there are distinct differences between them. In the three largest series of adult ALL,25;26;98 the proportion of cytogenetically normal cases was consistently higher in T-ALL than in B-lineage ALL (range of 24–43% versus 13–32%, respectively). About one-third of T-ALL have a translocation involving one of the T-cell receptor genes (TCR), resulting in over-expression of a gene juxtaposed to a TCR (Table 5). A breakpoint in 14q11.2, the location of a-TCR and d-TCR, is the
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Table 5 Breakpoints and genes recurrently rearranged with T-cell receptor genes in T-ALL Chromosome breakpoint
Gene involved
1p32 1p35-p34.3 8q24 9q34.3 10q24 11p13 11p15.5 19p13.2-p13.1
SCL (TAL-1 or TCL-5) LCK PVT1 and cMYC TAN-1 HOX11 RBTN2 (TTG2) RBTN1 (TTG1) LYL1
most common TCR breakpoint, but breakpoints in 7q35 (b-TCR) and 7p15 (c-TCR) also recur. The most common TCR rearrangement in adult ALL is t(10;14)(q24;q11.2), which results in overexpression of the HOX11 gene.98 It appears to be a primary aberration and is associated with a favorable outcome.98 Few studies of a recurrent d-TCR rearrangement in children, t(1;14)(p32;q11.2), have been reported in adults.31 This translocation juxtaposes d-TCR and SCL (also called TAL1 or TCL5), resulting in over-expression of SCL. A cryptic deletion of chromosome 1 in which SCL is juxtaposed to SIL also results in overexpression of SCL. In pediatric T-ALL, t(1;14) occurs only in about 3% of cases, but the SIL-SCL deletion occurs in 6–26%, making the combination of the two rearrangements the most common abnormality in pediatric TALL.157 The frequency of these rearrangements in adult ALL has not been widely studied. A cryptic t(5;14)(q35;q32) occurs in 20–30% of pediatric T-ALL;158;159 it is less common in adults. The translocation results in over-expression of HOX11L2 (5q35), although the breakpoint is actually in RanBP17, distant from HOX11L2. The abnormal expression of HOX11L2 is thought to result from abnormal control of the gene by CPT12, located at 14q32.158 Translocation (4;11)(q21;p15.5) is a rare abnormality that occurs in T-ALL,160;161 frequently in association with a del(12p). It is more frequent in children and young adults, thus far with poor outcomes. The rearrangement fuses NUP98 from chromosome 11 with RAP1GDS1 from chromosome 4 to encode nrg, a fusion protein.160;161
Concluding remarks During the last 30 years, cytogenetic analyses of patients with AML and ALL have discovered a great number of recurrent chromosome abnormalities. Several of the more common abnormalities have
been associated with specific laboratory and clinical characteristics, and are being used as diagnostic and prognostic markers that can guide the clinician in selecting the most effective treatment regimens. However, the prognostic importance of less frequent recurrent aberrations, both primary and secondary, is still unknown. Continuing cytogenetic studies are thus necessary to accrue enough patients with these rarer abnormalities to define conclusively their impact on CR rates, remission duration and survival, and to resolve discrepancies in prognostic categorization of some of the more frequent aberrations that currently exist among the major cytogenetic risk-assignment systems. Such studies will likely uncover new recurrent aberrations as they are still being identified by both classical cytogenetic methods and, increasingly, by molecular-cytogenetic techniques such as FISH, spectral karyotyping (SKY), multiplex-FISH (M-FISH) and multiplex FISH telomere assay (MTEL).38;39;68;71;158;159;162–165 Moreover, it is well known that prognostic factors depend on the type of therapy used. A cytogenetic or molecular genetic abnormality conferring an adverse prognosis with one therapeutic regimen may lose its unfavorable prognostic impact when another treatment is used. Therefore there is a constant need for large prospective studies correlating karyotype with selected molecular genetic markers, gene expression profiles, immunophenotype, other biologic parameters and clinical outcome in patients treated with both current therapies and those receiving novel therapeutic agents.
Practice points • Cytogenetic analysis should be performed in all newly diagnosed patients with AML or ALL. • Patients may be stratified to different therapies based on results of standard cytogenetic analysis, FISH and/or molecular genetic investigations (RT-PCR).
Research agenda • Continuing correlation of recurrent chromosome aberrations with response rates, response duration, survival and cure in groups of AML and ALL patients treated with current and novel induction and post-induction regimens. • Ascertainment of prognostic significance of the less common recurrent abnormalities in AML and ALL that are currently arbitrarily assigned to the intermediate-risk group or not categorized at all.
Cytogenetics in acute leukemia
• Further correlation of specific cytogenetic findings with biological and clinical features of acute leukemias to define unique disease entities that will be incorporated into the WHO Classification of Tumors of Hematopoietic and Lymphoid Tissues in the future. • Molecular genetic detection of submicroscopic mutations in patients belonging to cytogenetically defined groups (e.g., normal karyotype) that may provide additional clinically relevant information.
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Acknowledgements 15.
This work was supported in part by grant 5P30CA16058 from the National Cancer Institute, Bethesda, MD, and the Coleman Leukemia Research Fund.
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169. Raimondi SC, Chang MN, Ravindranath Y, et al. Chromosomal abnormalities in 478 children with acute myeloid leukemia: clinical characteristics and treatment outcome in a cooperative Pediatric Oncology Group study – POG 8821. Blood 1999;94:3707–16. 170. Larson RA, Dodge RK, Burns CP, et al. A five-drug remission induction regimen with intensive consolidation for adults with acute lymphoblastic leukemia: Cancer and Leukemia Group B study 8811. Blood 1995;85:2025–37. 171. Uckun FM, Nachman JB, Sather HN, et al. Poor treatment outcome of Philadelphia chromosome-positive pediatric acute lymphoblastic leukemia despite intensive chemotherapy. Leuk Lymphoma 1999;33:101–6. M, Valsecchi MG, Camitta B, et al. Outcome of 172. Arico treatment in children with Philadelphia chromosomepositive acute lymphoblastic leukemia. N Engl J Med 2000;342:998–1006. 173. Pui C-H, Gaynon PS, Boyett JM, et al. Outcome of treatment in childhood acute lymphoblastic leukaemia with rearrangements of the 11q23 chromosomal region. Lancet 2002;359:1909–15. 174. Uckun FM, Sensel MG, Sather HN, et al. Clinical significance of translocation t(1;19) in childhood acute lymphoblastic leukemia in the context of contemporary therapies: a report from the Children’s Cancer Group. J Clin Oncol 1998;16:527–35. 175. Crist WM, Carroll AJ, Shuster JJ, et al. Poor prognosis of children with pre-B acute lymphoblastic leukemia is associated with the t(1;19)(q23;p13): a Pediatric Oncology Group study. Blood 1990;76:117–22. 176. Heerema NA, Sather HN, Sensel MG, et al. Association of chromosome arm 9p abnormalities with adverse risk in childhood acute lymphoblastic leukemia: a report from the Children’s Cancer Group. Blood 1999;94:1537–44. 177. Heerema NA, Sather HN, Sensel MG, et al. Prognostic significance of cytogenetic abnormalities of chromosome arm 12p in childhood acute lymphoblastic leukemia: a report from the Children’s Cancer Group. Cancer 2000;88: 1945–54. 178. Heerema NA, Sather HN, Sensel MG, et al. Clinical significance of deletions of chromosome arm 6q in childhood acute lymphoblastic leukemia: a report from the Children’s Cancer Group. Leuk Lymphoma 2000;36: 467–78. 179. Rao KW, Heerema NA, Sather HN, et al. Deletions of chromosome arm 5q in childhood acute lymphoblastic leukemia (ALL). Proc Am Soc Clin Oncol 2000;19:589a. 180. Raimondi SC, Behm FG, Roberson PK, et al. Cytogenetics of childhood T-cell leukemia. Blood 1988;72:1560–6. 181. Heerema NA, Sather HN, Sensel MG, et al. Frequency and clinical significance of cytogenetic abnormalities in pediatric T-lineage acute lymphoblastic leukemia: a report from the Children’s Cancer Group. J Clin Oncol 1998;16:1270–8.
www.elsevierhealth.com/journals/blre
Cytogenetics in acute leukemia zeka,*, Nyla A. Heeremab, Clara D. Bloomfielda Krzysztof Mro a
Division of Hematology and Oncology, The Comprehensive Cancer Center, The Arthur G. James Cancer Hospital and Richard J. Solove Research Institute, Room 1248B, The Ohio State University, 300 West Tenth Avenue, Columbus, OH 43210-1228, USA b Department of Pathology, The Comprehensive Cancer Center, The Ohio State University, Columbus, OH 43210-1228, USA
KEYWORDS Acute myeloid leukemia; Acute lymphoblastic leukemia; Chromosome aberrations; Human; Karyotyping; Prognosis
Summary Cytogenetic analyses in acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) have revealed a great number of non-random chromosome abnormalities. In many instances, molecular studies of these abnormalities identified specific genes implicated in the process of leukemogenesis. The more common chromosome aberrations have been associated with specific laboratory and clinical characteristics, and are now being used as diagnostic and prognostic markers guiding the clinician in selecting the most effective therapies. Specific chromosome aberrations and their molecular counterparts have been included in the World Health Organization classification of hematologic malignancies, and together with morphology, immunophenotype and clinical features are used to define distinct disease entities. However, the prognostic importance of less frequent recurrent aberrations in AML and ALL, both primary and secondary, is still to be determined. This review summarizes current views on clinical relevance of major cytogenetic findings in adult AML and ALL. c 2003 Elsevier Ltd. All rights reserved.
Introduction The role of cytogenetics in determining the biologic basis of acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) is now widely recognized. By identifying acquired chromosome aberrations that recur in AML and ALL, and providing precise chromosomal location of breakpoints in leukemia-associated translocations and inversions, cytogenetics aided in cloning of many genes whose activation or fusion with other genes contributes to the neoplastic process.1–3 Further
*
Corresponding author. Tel.: +1-614-293-3150; fax: +1-614293-3457. zek). E-mail address: [email protected] (K. Mro
characterization of these genes revealed that they are often involved directly or indirectly in the development and homeostasis of normal blood cells, and that abnormal protein products of fusion genes created by specific translocations and inversions can dysregulate proliferation, differentiation or programmed cell death (apoptosis) of blood cell precursors.1 This has paved the way to designing novel therapeutic agents targeting specific genetic abnormalities in leukemic blasts, such as imatinib mesylate, the Bcr–Abl tyrosine kinase inhibitor that suppresses proliferation of cells harboring the BCR–ABL fusion gene created by t(9;22) (q34;q11.2), a recurrent chromosome aberration in chronic myelogenous leukemia (CML) and ALL.4 Furthermore, cytogenetic analysis is now routine part of clinical practice, being an important
0268-960X/$ - see front matter c 2003 Elsevier Ltd. All rights reserved. doi:10.1016/S0268-960X(03)00040-7
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tool in diagnosis and prognostication of acute leukemias. Specific chromosome aberrations and their molecular counterparts have been included in the World Health Organization (WHO) classification of hematologic malignancies, and together with morphology, immunophenotype and clinical features are being used to define distinct disease entities.5–7 Pretreatment cytogenetic findings have been repeatedly shown to be among the most important, independent prognostic factors in both AML8–20 and ALL.21–26 Consequently, cytogenetic analyses are considered mandatory for analyzing outcome of many clinical trials and are currently used to stratify patients for different types of therapy.27–29 In this review, we will summarize the clinical relevance of major cytogenetic findings in adult AML and ALL.
Acute myeloid leukemia Clonal chromosome abnormalities, that is, an identical structural aberration or gain of the same, structurally intact chromosome detected in at least two metaphase cells or the same chromosome missing from a minimum of three cells, are consistently found in the majority of AML patients at diagnosis. However, in contrast to patients diagnosed with CML, who are invariably positive for t(9;22) or its variants, the cytogenetic picture of AML is much more complex. To date, approximately 200 different structural and numerical aberrations such as reciprocal translocations, inversions, insertions, deletions, unbalanced translocations, isochromosomes, isodicentric chromosomes, isolated trisomies and monosomies have been found to be recurring chromosome changes in AML. Many of these aberrations are very rare, being so far detected in a few patients worldwide, whereas others occur more frequently. These more common abnormalities, together with their frequencies among adults and children with AML, are presented in Table 1. A more complete list of specific AML-associated recurrent chromosome aberrations, and genes affected by their formation (if known), can be found in Refs. 27, 30–32. The incidence of abnormal karyotypes is in general lower in adult than in pediatric de novo AML. While 55% of 4257 adults with AML enrolled onto three large cooperative studies displayed chromosome aberrations (range 53–60%), 76% of 1184 patients included in the four largest series of childhood AML had an abnormal karyotype (range 68–85%; Table 1). The reasons for such a discrep-
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ancy are unknown, but may be related to biological differences between pediatric and adult disease. This is likely reflected by varying proportions of specific chromosome aberrations in children and adults with AML. For example, balanced rearrangements, mostly reciprocal translocations, involving band 11q23 and disrupting the MLL gene are on average four times more common in children than in adults. The frequency of 11q23 rearrangements in patients with AML decreases appreciably with age: they are detected in 43–58% of infants aged 12 months or less,33–35 in 39% of children aged 13–24 months,35 in 8–9% of children older than 24 months,35 and in just 4–7% of adults (Table 1), among whom, according to the United Kingdom Medical Research Council (MRC) study,19 the frequency of 11q23 rearrangements goes down from 5% in patients aged 15–34 years to 2% among those aged 35–55 years and 1% in patients older than 55 years. Some other, relatively rare, recurrent aberrations are essentially restricted to young children with AML, not being detected in adult patients thus far. The t(1;22)(p13;q13), a translocation resulting in the OTT-MAL gene fusion and highly correlated with acute megakaryoblastic leukemia, has been to date detected exclusively in children, 96% of whom were younger than 24 months.36;37 Interestingly, almost all infants younger than 6 months had t(1;22) as a sole aberration whereas it was predominantly part of a complex karyotype in the majority of older children (p ¼ 0:00004).36 Another aberration hitherto detected only in infants aged 20 months or younger is t(7;12) (q36;p13). This subtle translocation, involving the ETV6 gene and almost always occurring together with trisomy 19, was often misdiagnosed as del(12p) in the past and only recently has been shown using fluorescence in situ hybridization (FISH) to be a consistent chromosome aberration in infant AML.38;39 In contrast, t(15;17)(q22;q12-21) and t(8;21)(q22;q22), the two most common reciprocal translocations in both adults and older children with AML (Table 1), have never been detected in infants aged less than 12 months.31 However, while the incidence of t(15;17) [and inv(16)(p13q22)/t(16;16)(p13;q22)] is similar in adults and older children, t(8;21) is twice as common in pediatric as in adult AML (Table 1). On the other hand, )5, del(5q) and other unbalanced structural abnormalities resulting in loss of material from 5q are much more frequent in adult than childhood AML. Likewise, both inv(3)(q21q26) and t(3;3)(q21;q26), which are found in 2% of adults, are extremely rare in children and have so far never been detected in a patient with de novo AML younger than 12 years.31
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Table 1 Frequencies of the more common cytogenetic abnormalities in adult and childhood AML Cytogenetic abnormality
None (normal karyotype) +8 )7/7q) )7 del(7q) Loss of (7q)f t(15;17)(q22;q21) )5/5q) )5 del(5q) Loss of (5q)f t(8;21)(q22;q22) inv(16)(p13q22)/ t(16;16)(p13;q22) )Y t/inv(11q23) t(9;11)(p22;q23) abn(12p) +21 abn(17p) del(9q) inv(3)(q21q26)/ t(3;3)(q21;q26) del(11q) t(9;22)(q34;q11) t(6;9)(p23;q34) Complex karyotype with P3 abn Complex karyotype with P5 abn
Cooperative Group Study (No. of patients) CALGBc ðn ¼ 1311Þ No. (%)
MRCd ðn ¼ 2337Þ No. (%)
SWOG/ECOGe ðn ¼ 609Þ No. (%)
Adults totala ðn ¼ 4257Þ No. (%)
Childrena;b ðn ¼ 1184Þ No. (%)
582 (44)
1096 (47)
244 (40)
1922 (45.1)
283 (23.9)
123 (9) 95 (7) 47 (4) 19 (1) 29 (2) 88 (7) 86 (7) 26 (2) 42 (3) 18 (1) 81 (6) 96 (7)
211 (9) 209 (9) 136 (6) 73 (3) NA 210 (9) 183 (8) 79 (3) 104 (4) NA 104 (4) 53 (2)
53 (9) 52 (9) NA NA NA 27 (4) 36 (6) NA NA NA 50 (8) 53 (9)
387 (9.1) 356 (8.4) 183 (5.0) 92 (2.5) 29 (2.2) 325 (7.6) 305 (7.2) 105 (2.9) 146 (4.0) 18 (1.4) 235 (5.5) 202 (4.7)
112 (9.5) 62 (5.2) 33 (2.8) 18 (1.5) 11 (1.3) 117 (9.9) 14 (1.2) 4 (0.3) 9 (0.8) 1 (0.1) 137 (11.6) 70 (5.9)
58 54 27 33 28 30 33 12
(4) (4) (2) (3) (2) (3) (3) (1)
NA 45 (2) NA NA 51 (2) NA 37 (2) 61 (3)i
20 (3) 42 (7)g NA NA NA 12 (2)h 17 (3) 12 (2)
78 (4.1) 141 (3.3) 27 (2.1) 33 (2.5) 79 (2.2) 42 (2.2) 87 (2.1) 85 (2.0)
28 (3.8) 155 (13.1) 54 (6.4) NA 60 (5.1) 5 (2.0) 33 (2.8) 0
12 (1) 10 (1) 8 (1) 135 (10)
NA 16 (1) 10 (<1) NA
NA 8 (1) 11 (2) 71 (12)
12 (0.9) 34 (0.8) 29 (0.7) 206 (10.7)
11 (1.3) 2 (0.2) 12 (1.0) 36 (14.3)
99 (8)
222 (9)
53 (9)
374 (8.8)
31 (5.2)
Abbreviations: CALGB, Cancer and Leukemia Group B; MRC, United Kingdom Medical Research Council; SWOG/ ECOG Southwest Oncology Group/Eastern Cooperative Oncology Group; abn, abnormality; NA, not available. a Percentages for particular abnormalities calculated using only those studies that provided relevant data. b Data from Leverger et al.166 (130 children aged from 2 months to 16 years diagnosed with untreated AML); Raimondi et al.167 (121 children diagnosed with de novo AML); Martinez-Climent et al.168 (115 children and adolescents aged from 0 months to 19.2 years diagnosed with de novo AML); Raimondi et al.169 (478 children and adolescents younger than 21 years diagnosed with de novo AML). c Data from Byrd et al.;20 the study comprised patients aged 15–86 years diagnosed with de novo AML. d Data from Grimwade et al.16 and Grimwade et al.;19 Grimwade et al.16 comprised 1272 patients aged 15–55 years, the majority of whom were diagnosed with de novo AML; up to 9.4% of patients had AML secondary to chemotherapy or radiotherapy or to an antecedent hematologic disorder. Grimwade et al.19 comprised 1065 patients aged 44–91 years, 817 of whom were diagnosed with de novo AML and 248 with AML secondary to chemotherapy or radiotherapy or to an antecedent hematologic disorder. e Data from Slovak et al.;18 the study comprised patients aged 16–55 years diagnosed with untreated AML. f Loss of 7q and Loss of 5q refer to unbalanced structural abnormalities, other than del(7q) and del(5q), that result in loss of material from the, respectively, 7q and 5q chromosome arms (e.g., unbalanced translocations, additions, isochromosome of 7p or 5p, etc.). g Category defined as “abn 11q”, might also include patients with other abnormalities of 11q including del(11q). h Including three cases with i(17)(q10). i Category defined as “abn 3q”, might also include patients with other abnormalities of 3q.
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Prognostic significance of cytogenetics in AML The Fourth International Workshop on Chromosomes in Leukemia (4IWCL) was the first large, prospective, multi-center study that established pretreatment karyotype as an independent prognostic factor in AML.8 Significant differences in complete remission (CR) rate, CR duration (CRD) and overall survival (OS) were demonstrated when the 716 patients were prioritized first according to the presence of t(8;21); then t(15;17); followed by )5 or del(5q), )7 or del(7q); concurrent occurrence of )5 or del(5q) and )7 or del(7q), and then abnormalities of 11q, +8 and +21. Patients without any of the above-mentioned abnormalities were classified according to the chromosome number [hypodiploid, pseudodiploid, diploid (normal) and hyperdiploid]. Cytogenetic findings so classified were independent prognostic factors for both duration of first CR and overall survival in the subset of 305 patients treated adequately.8 In the followup studies, a group of patients with abnormalities of 16q22 was added, all cases with aberrations of chromosomes 5 and 7 were combined into one category, and patients with +8 and +21 included in the hyperdiploid group.12;14;40 The multivariate analyses performed at the third follow-up of the 4IWCL, which then comprised 628 cases with de novo AML with a median follow-up of 14.7 years for patients alive, confirmed karyotype as an independent predictor of survival for all patients and for those 291 patients who received induction therapy that would be deemed standard by present-day criteria.14 Many studies, both smaller, performed in a single institution and large, collaborative multi-institutional ones, have confirmed that pretreatment karyotype constitutes an independent prognostic determinant for attainment of CR, CRD, risk of relapse and survival.9–11;13;15;16;18;20 Recently, three large collaborative studies proposed prioritization schemata that assign AML patients to one of the three risk groups, favorable, intermediate or adverse, based on pretreatment cytogenetic findings.16;18;20 The three cytogenetic risk systems share many common features, but differ with regard to some aspects (Table 2). In the MRC schema, any abnormality that is not classified as favorable or adverse, and is not accompanied by any additional chromosome aberration classified as favorable or adverse, is categorized in the intermediate-risk group. In contrast, both the Southwest Oncology Group/Eastern Cooperative Oncology Group (SWOG/ECOG) and Cancer and Leukemia Group B (CALGB) schemata explicitly
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categorize particular abnormalities into risk groups, and leave as not classified aberrations too infrequent to be analyzed. Moreover, MRC and SWOG/ECOG classify patients with a given abnormality into one of the three risk groups once, whereas CALGB provides risk-group assignment separately for probability of induction success, cumulative incidence of relapse (CIR) and OS. Consequently, in the CALGB schema, the same abnormality [e.g., t(6;9)(p23;q34)] may be categorized in the intermediate-risk group with regard to probability of achieving a CR, in the adverse-risk group with respect to OS and not being classified with regard to CIR, because too few patients attained a CR to be analyzed. Despite these differences, the inv(16)/t(16;16) and t(8;21), cytogenetic hallmarks of core-binding factor (CBF) AML, have been categorized in the favorable group by all three cytogenetic risk systems. However, SWOG/ECOG classified as favorable also patients with del(16q) whereas MRC and CALGB did not. We believe that patients with true del(16q) [i.e., deletions that do not represent misinterpreted inv(16) or t(16;16) and are usually found in AML with morphology other than that of acute myelomonocytic leukemia with abnormal eosinophils (AMML Eo)] should not be included in the favorable risk group, because del(16q) differs from inv(16)/t(16;16) at the molecular level, and has not been associated with a favorable outcome comparable to that of inv(16)/t(16;16).28;41;42 Consequently, the RT-PCR and/or FISH assays detecting CBFb-MYH11 gene fusion should be performed in all patients with del(16)(q22) to ensure that they do not harbor a misidentified inv(16)/ t(16;16).28 Moreover, while MRC and CALGB included all patients with t(8;21) in the favorable group, irrespective of whether t(8;21) was the sole aberration or occurred together with one or more secondary abnormalities, SWOG/ECOG classified as favorable only those t(8;21)-positive patients who did not have a complex karyotype with three or more abnormalities or a secondary del(9q). The latter abnormality has been reported as a poor risk indicator in patients with t(8;21) in one study,43 but this result was not corroborated by the MRC,16 CALGB,20 nor by other studies.8;10;44 Indeed, it has been consistently reported that for patients with t(8;21) and inv(16)/t(16;16), neither the presence of secondary abnormalities, including the most frequent in t(8;21)-positive patients )Y, )X and +8, nor a complex karyotype with three or more abnormalities adversely affects clinical outcome.8;10;16;20;44 Instead, adverse prognostic significance among t(8;21)-positive patients has been attributed to such factors as high initial white
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Table 2 Risk-group assignments of patients with selected chromosome aberrations in the 3 major collaborative cytogenetic studies of adult AML
Abbreviations: CALGB, Cancer and Leukemia Group B; MRC, United Kingdom Medical Research Council; SWOG/ ECOG Southwest Oncology Group/Eastern Cooperative Oncology Group; CR, complete remission; CIR, cumulative incidence of relapse; OS, 5-year overall survival; abn, abnormality; NA, not available. a For the convenience of the reader, risk group assignments of particular abnormalities are depicted in color: green for “favorable”, blue for “intermediate” and probably intermediate by virtue of being “other structural” or “other numerical” in the MRC classification, red for “adverse” and brown for “not classified” or “unknown”.
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Table 2 (continued) b Data from Byrd et al.;20 1213 adults with de novo AML analyzed. Patients with t(15;17) and t(9;22) were excluded from the analysis because therapy with ATRA or imatinib mesylate, respectively, is now considered a standard treatment option for these AML subtypes. All patients with t(8;21) and inv(16)/t(16;16) were classified in the favorable and those with t(9;11) in the intermediate risk categories regardless of whether t(8;21), inv(16)/t(16;16) and t(9;11) occurred alone or together with any secondary aberration or as part of a complex karyotype (defined as the presence of three or more aberrations). All other patients with a complex karyotype were classified as having adverse risk. Consequently, risk-group assignments of all other aberrations concerned only those patients who had a given aberration occurring alone or with only one additional abnormality other than t(8;21), inv(16)/t(16;16) and t(9;11).20 c Data from Grimwade et al.16 Of the 1612 patients analyzed, 1493 were diagnosed with de novo AML and 119 with secondary AML. The patient population included 340 children aged 0–14 years and 1272 adults aged 15–55 years. Patients with t(8;21), inv(16)/t(16;16) and t(15;17) were classified in the favorable risk group irrespective of whether t(8;21), inv(16)/t(16;16) and t(15;17) occurred alone or in conjunction with other abnormalities. Adverse abnormalities were classified as such regardless of whether they occurred alone or in conjunction with intermediate-risk or other adverse-risk abnormalities. Intermediate group comprised cytogenetic abnormalities not classified as favorable or adverse in the absence of accompanying favorable or adverse cytogenetic changes16 . d Data from Slovak et al.;18 609 adults aged 16–55 years with untreated AML analyzed. e Provided t(8;21) is not part of a complex karyotype or is not accompanied by del(9q). Both complex karyotype and del(9q) were classified in the adverse prognostic category. f Favorable for a group of 13 patients with del(9q) that included six who underwent SCT off-protocol; intermediate for non-transplanted patients treated with chemotherapy only. g Would be included in “abnormal 11q23” category. h Would be included in “abn 20q” category. i del(12p) classified as intermediate, other rearrangements affecting 12p presumably classified as unknown. j The abnormality accompanying +8 is other than t(8;21), t(9;11) and inv(16) or t(16;16). k Intermediate provided the abnormality accompanying +8 is not classified as favorable or adverse. l Would be included in “abn 3q” category. m Would be included in “abn 11q” category.
blood cell count (WBC) or absolute granulocyte count, the presence of granulocytic sarcomas, expression of the neural cell adhesion molecule CD56 on leukemic blasts and high WBC index.44 The WBC index [i.e., WBC (% of marrow blasts/100)], which divides patients into three subgroups (low index, <2.5; intermediate index, 2.5–20; high index, P 20) that differ with respect to clinical outcome, has been found in a recent large study to be the only prognostic factor for CRD, disease-free survival (DFS) and overall survival of patients with t(8;21) in multivariate analysis.44 The outcome of adults with CBF AML can be improved substantially by intensive post-remission therapy with high-doses of cytarabine (HDAC).17 Furthermore, patients with t(8;21) who received three or four cycles of HDAC on CALGB treatment protocols had superior projected 5-year DFS (71% versus 37%) and survival (76% versus 44%) rates compared with those who were administered only one cycle of HDAC followed by sequential treatments with cyclophosphamide/etoposide and mitoxantrone/diaziquone with or without filgrastim.45 The efficacy of repeated courses of HDAC for AML patients with t(8;21) has been confirmed by others.46 Likewise, the 5-year CIR was signifi-
cantly decreased in patients with inv(16)/t(16;16) receiving 3–4 cycles of HDAC as compared with those receiving one HDAC course (43% versus 70%) [Byrd et al., unpublished results]. This improved outcome is likely related to an increased sensitivity of the leukemic cells carrying inv(16)/t(16;16) or t(8;21) to cytarabine. Tosi et al.47 have shown a significant increase in incorporation of cytarabine into nuclear DNA in vitro and cytarabine induced apoptosis in blasts from patients with inv(16) compared to cells from patients with other chromosome aberrations or a normal karyotype. Another study revealed that samples from patients with inv(16)/t(16;16) and those with t(8;21) had the highest in vitro proliferative activity which was closely correlated with an increased incorporation of the active moiety of cytarabine, AraC triphosphate, into the DNA.48 The three major cytogenetic risk classifications agree that prognosis of patients with inv(3) (q21q26) or t(3;3)(q21;q26), )7 and a complex karyotype is poor (Table 2). The definition of complex karyotype differs, though, with the MRC characterizing this category as “the presence of a clone with at least five unrelated cytogenetic abnormalities”, and SWOG/ECOG and CALGB, as well
Cytogenetics in acute leukemia
as the German AML Study Group,49 as three or more abnormalities. A multi-center Italian study defined complex karyotype as “the presence of a clone with more than three cytogenetic abnormalities”.50 Byrd et al.20 compared the outcome of patients with three or four abnormalities [other than t(8;21), inv(16)/t(16;16) or t(9;11)(p22;q23)] with that of patients with five or more abnormalities. Although patients in the former group were younger and had significantly better CIR and 5-year OS than patients with five or more abnormalities, the CR rate and OS of patients with three or four abnormalities were significantly lower and the CIR significantly higher than those of patients included in the cytogenetically normal group. Only one patient in the group with three or four abnormalities remained in remission at 5 years. Thus, the authors concluded that these data justify combining patients with three or four abnormalities with patients who have five or more abnormalities into one complex karyotype category defined by the presence of three or more abnormalities.20 It has been striking in the CALGB series that all or almost all patients with )5, del(7q), )17/17p), )18, and )20 had a complex karyotype, thus precluding assessment of the prognostic significance of these abnormalities independently from complex karyotype. In the MRC study, the outcome of patients with del(7q) that was not part of a complex karyotype with P 5 abnormalities and was not accompanied by )5/del(5q) or abn(3q) did not differ significantly from outcome of patients with a normal karyotype.16 Their findings are consistent with earlier observations suggesting that patients with del(7q) without coexisting aberrations of chromosome 5 may have prolonged survival.40;51 In the CALGB series, most patients with del(5q) had a complex karyotype and very poor prognosis. However, a relatively small group of patients with del(5q) in a non-complex karyotype was classified as having intermediate risk with regard to CR rate and OS.20 Patients with various balanced abnormalities involving band 11q23 have been grouped into one cytogenetic category in both the SWOG/ECOG and MRC study, and classified, respectively, as having adverse and intermediate prognosis (Table 2). However, mounting evidence suggests that outcome of patients with translocations involving band 11q23 depends on the partner chromosome involved, with t(9;11)(p22;q23)-positive patients having a more favorable prognosis that places them in the intermediate-risk group.20;52 Remarkably, a recent study of 298 infants and children with de novo AML found that t(9;11) was the most important, favorable prognostic factor for patients
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treated on protocols used at St. Jude Children’s Research Hospital.53 Zwaan et al.54 suggested that the superior outcome of the t(9;11)-positive patients may be explained by enhanced sensitivity of their leukemic blasts to several chemotherapeutic drugs. In this study, bone marrow or blood samples from children with AML and t(9;11) were significantly more sensitive in vitro to cytarabine, etoposide, the anthracyclines and 2-chlorodeoxyadenosine than samples from patients with various other chromosome aberrations. When compared with a subgroup of childhood AML patients harboring other 11q23 translocations, including t(6;11)(q27;q23), t(9;11)-positive samples were significantly more sensitive for cytarabine and doxorubicin, and borderline more sensitive for etoposide.54 The survival of adults with t(6;11) and t(11;19)(q23;p13.1) studied by CALGB was significantly shorter than that of the cytogenetically normal group, and, consequently, t(6;11) and t(11;19)(q23;p13.1) were assigned to the adverserisk group for OS. For patients with other, less frequent 11q23 translocations, the definitive assignment of risk category will be possible only once enough patients are analyzed in large prospective studies.20 The most common trisomies in de novo AML are, in decreasing order of frequency, +8, +22, +13, +21 and +11.20 Trisomy 22 is a non-random secondary aberration accompanying inv(16)/t(16;16) and is rarely seen as the only chromosome abnormality.31 Although each of the remaining trisomies can be found as a secondary aberration, +8, +13, +11 and +21 are also detected recurrently as the only (isolated) karyotypic changes at diagnosis, with a frequency among adults with de novo AML of 4% for sole +8, 1% each for sole +13 and +11, and 0.4% for sole +21.55 With regard to the impact of recurrent trisomies on clinical outcome, most data have been gathered for trisomy 8, but results have been somewhat inconsistent. CR rates of patients with +8 have differed widely, from 29%11 to 91%,51 as have CRD and survival among studies,32 and consequently patients with +8 have been classified either in the intermediate or adverse-risk category. In some reports, this cytogenetic group included both patients with isolated +8 and those who in addition to +8 had other aberrations that may have affected response to treatment and outcome. It has been repeatedly shown that prognosis of AML patients with +8 indeed depends on whether +8 occurs as an isolated abnormality or is accompanying other aberrations.16;20;56–58 In the latter situation, +8 does not appear to adversely affect the favorable outcome of patients with t(15;17), inv(16)/t(16;16) and t(8;21).16;20;56;57 In contrast,
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patients with +8 and a complex karyotype and/or such unfavorable aberrations as del(5q) or )7 usually have very poor outcome. In one recent study, patients with +8 and unfavorable chromosome aberrations (according to the SWOG/ECOG classification) had significantly worse OS than other patients with unfavorable aberrations who lacked +8 in their karyotype.58 On the other hand, isolated +8 has been considered to bestow either intermediate or unfavorable prognosis (Table 2). The conflicting results might be associated with variation in the age of the patients analyzed in different series,30 but may also stem from differences in proportion of patients who underwent stem cell transplantation (SCT). Farag et al.55 reported that prognosis of patients with an isolated +8, +11, +13 and +21 was dependent on SCT. While these isolated trisomies constituted an independent adverse prognostic factor for OS in non-transplanted patients, in those receiving SCT in first remission, the 5-year OS was not different than that of transplanted patients with a normal karyotype thus changing the risk stratification of isolated trisomies from adverse to intermediate.55 Prospective studies are required to confirm this finding. Patients with a normal karyotype are the single largest cytogenetic subset of adult AML. They are classified in the intermediate prognostic category, because their CR rates, CRD and survival probabilities are usually lower than those of adequately treated patients with t(8;21), inv(16) or t(15;17), but higher than patients with unfavorable chromosome aberrations.16;20;51 However, this group is highly heterogeneous at the molecular level, and is likely composed of subsets with varying prognoses. A small fraction of patients determined to have a normal karyotype by conventional cytogenetic analysis may carry gene fusions typically associated with recurrent translocations or inversions. Occasionally, this happens because the presence of a chromosome aberration is not recognized by the cytogenetic laboratory, especially when the quality of chromosome preparation is suboptimal and the missed rearrangement subtle, e.g., t(11;19)(q23;p13.1), t(15;17), or inv(16). However, in other patients, the gene fusion results from a cryptic rearrangement involving segments smaller than the length of a single band and thus unrecognizable by standard cytogenetic analysis. For instance, cryptic insertions of a very small segment from 17q containing the RARa gene into the locus of the PML gene on chromosome 15q have been repeatedly detected using FISH in patients with acute promyelocytic leukemia (APL) and a normal karyotype.59 Grimwade et al.59 estimates that insertions leading to formation of the PML-RARa fu-
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sion gene, most of which are cryptic, account for approximately 4% of all patients with APL. Because marrow of patients with cryptic PML-RARa fusions has a typical APL morphology, and such patients share the favorable prognosis of those with the standard t(15;17),59 karyotypically normal patients suspected of having APL should be tested for the presence of the PML-RARa fusion gene by reverse transcription-polymerase chain reaction (RT-PCR), Southern blot analysis or/and FISH, and, if found positive, treated with regimens containing alltrans-retinoic acid (ATRA). Similarly, rare patients with the CBFb-MYH11 fusion gene but without microscopically detectable abnormalities of chromosome 1660;61 and those with the RUNX1-CBFA2T1 (AML1-ETO) fusion gene and normal chromosomes 8 and 2162;63 should likely be grouped together with other patients with CBF AML. Of note, as demonstrated by two large prospective series,28;63 patients truly positive for the CBFb-MYH 11 and Runx1-CBFA2T1 gene fusion transcripts in the absence of cytogenetically detectable aberrations of chromosomes 16, and 8 and 21, respectively, are rare and their frequencies are much lower than those indicated by two retrospective studies.64;65 . It is also rather unlikely that a large proportion of cytogenetically normal patients with de novo AML harbor hidden aberrations detectable only by FISH-based techniques. In a series of 102 adult AML patients tested by FISH using a comprehensive set of genomic DNA probes capable of detecting major AML-associated aberrations, only three patients were found to harbor, respectively, inv(16), )Y and del(12p).66 In retrospect, only the del(12p) appeared to represent a truly cryptic aberration.66 Cuneo et al.67 did not find any occult chromosome anomaly among 55 adults with de novo AML aged 15–60 years who were studied by FISH using probes detecting )5, )7, +8, and deletions of 5q31, 7q31, 12p13/ETV6, 17p13/TP53 and 20q11. The authors did identify, however, chromosome aberrations in interphase cells of 3 of 21 (14%) AML patients aged 62–80 years and in 5 of 6 (83%) patients with AML secondary to an antecedent MDS. In four of these patients, FISH analysis of metaphase cells revealed submicroscopic del(5)(q31) (three cases) and del(7)(q31) (one case).67 To date, application of multi-color FISH-based techniques that use chromosome painting probes, such as spectral karyotyping (SKY) and multiplex FISH (M-FISH), to study AML patients with a normal karyotype has failed to discover any novel hidden rearrangement, akin to the cryptic t(12;21)(p13;q22) in childhood pre-B ALL, which, due to the juxtaposition of similarly banded regions, can be discerned microscopically only by FISH.68–71 On the other hand, a new recur-
Rearrangementa
Age group
Prognostic relevanceb
No. pts with/no. pts without rearrangement
Reference number
11/87 7/61 15/30
72
Normal karyotype PTD MLL
Adults
PTD MLL
Adults
PTD MLL
Adults 16–60 years
PTD MLL FLT3 ITD
Adults 16–60 years Adults 16–60 years
FLT3 Asp835
Adults 16–60 years
FLT3 ITD versus FLT3 Asp835 FLT3 ITD
Adults 16–60 years
FLT3 ITD
Adults 20–59 years
FLT3ITD=
Adults 20–59 years
FLT3ITD=WT
Adults 20–59 years
Adults
FLT3ITD= versus FLT3ITD=WT Adults 20–59 years
CR rate and OS: no significant difference CRD: significantly shorter for PTD MLL+ patients Median survival: significantly shorter for PTD MLL+ patients compared with that of age-matched karyotypically normal control group Relapse-free interval: significantly shorter for PTD MLL+ patients compared with that of age-matched karyotypically normal control group CR rate and OS: no significant difference CRD: significantly shorter for PTD MLL+ patients EFS: significantly shorter for PTD MLL+ patients CR rate: no significant difference CRD: significantly shorter for FLT3 ITD+ patients OS: significantly shorter for FLT3 ITD+ patients CR rate and OS: no significant difference CRD: no significant difference CR rate and OS: no significant difference CRD: no significant difference CR rate: no significant difference OS: significantly shorter for FLT3 ITD+ patients CRD: significantly shorter for FLT3 ITD+ patients CR rate and OS: no significant difference DFS: significantly shorter for FLT3 ITD+ patients CR rate: no significant difference OS: significantly shorter for FLT3ITD= patients DFS: significantly shorter for FLT3ITD= patients CR rate and OS: no significant difference DFS: no significant difference CR rate: no significant difference OS: significantly shorter for FLT3ITD= patients DFS: no significant difference
73
Cytogenetics in acute leukemia
Table 3 Molecular genetic rearrangements reported to affect clinical outcome of AML patients with a normal karyotype or those classified in the intermediate prognostic category
9/17
18/203 16/158 5/25 67/125 47/95 67/125 28/125 23/95 67/28 47/23 16/37 16/37 8/28 23/59 17/51 8/59 8/59 6/51 15/59 11/51 8/15 8/15 6/11
75 29 84
84 84 88
80 80
80 80
123
124
Table 3 (continued) Rearrangementa
Prognostic relevanceb
Age group
No. pts with/no. pts without rearrangement
Reference number
66/217 46/139 46/139 76/172 29/192
85
Intermediate-risk prognostic groupc FLT3 ITD
Adults <60 year
FLT3 ITD or FLT3 TKD Adults <60 year FLT3 ITD mutant/wt allele Adults <60 year ratio >0.78
FLT3 ITD mutant/wt allele Adults <60 year ratio >0 6 0.78 FLT3 ITD mutant/wt allele Adults <60 year ratio >0.78 versus >0 6 0.78
Adults
FLT3 ITD FLT3 ITD
Adults 15–74 years Adults
FLT3 ITD or FLT3 Asp835
Adults
FLT3 ITD FLT3 ITD N-RAS point mutation GSTM1 or/and GSTT1
Adults Adults Adults Adults
16–88 15–85 15–85 19–75
years years years years
85 85
23/152 23/152 30/192 25/152 25/152 29/30
85
85
23/25 23/25 118/288 109/251 44/94 10/34 15/65 17/54 27/65 26/54 15/40 123d 123d 51/44 31/36 51/44
86
77 89 89 78 76 76 94
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FLT3 ITD
CR rate: no significant difference DFS: no significant difference Probability of relapse: significantly higher for FLT3 ITD+ patients OS: significantly shorter for FLT3 ITD+/FLT3 TKD+ patients OS: significantly shorter for FLT3 ITD patients with mutant/ wt allele ratio >0.78 DFS: significantly shorter for FLT3 ITD patients with mutant/wt allele ratio >0.78 Probability of relapse: significantly higher for patients with mutant/wt allele ratio >0.78 OS: no significant difference DFS: no significant difference Probability of relapse: no significant difference OS: significantly shorter for FLT3 ITD patients with mutant/wt allele ratio >0.78 DFS: significantly shorter for FLT3 ITD patients with mutant/wt allele ratio >0.78 Probability of relapse: significantly higher for patients with mutant/wt allele ratio >0.78 EFS: significantly shorter for FLT3 ITD+ patients OS: no significant difference DFS: no significant difference OS: significantly shorter for FLT3 ITD+ patients EFS: no significant difference DFS: no significant difference EFS: no significant difference DFS: no significant difference DFS: significantly shorter for FLT3 ITD+ patients OS: significantly shorter for FLT3 ITD+ patients OS: significantly shorter for patients with N-RAS point mutation CR rate: significantly lower for GSTM1 /GSTT1 patients EFS: significantly shorter for GSTM1 /GSTT1 patients OS: significantly shorter for GSTM1 /GSTT1 patients
a
Unless otherwise indicated, all comparisons are between patients with a given gene rearrangement and those without (wild-type allele); PTD MLL, partial tandem duplication of the MLL gene; FLT3 ITD, internal tandem duplication of the FLT3 gene; FLT3ITD=WT cases of AML with the FLT3 ITD in combination with the FLT3 wildtype (WT) allele; FLT3ITD= , cases of AML with the FLT3 ITD lacking a FLT3 WT allele; FLT3 Asp835, activating point mutations of D835 within the activation loop of the FLT3 gene; FLT3 TKD, activating point mutations in codon 835 or 836 of the FLT3 gene; GSTM1 or/and GSTT1 , homozygous deletions resulting in null genotypes at the glutathione S-transferase GSTM1 or/and GSTT1 loci. b CR, complete remission; CRD, CR duration; EFS, event-free survival; OS, overall survival. c Intermediate-risk prognostic group defined as follows: Thiede et al.85 – normal karyotype and chromosome aberrations other than those conferring high risk [i.e., )5/del(5q),)7/del(7q), hypodiploid karyotypes (besides isolated )Y or )X), inv(3q), abn 12p, abn 11q, +11, +13, +21, +22, t(6;9); t(9;22); t(9;11); t(3;3) and multiple aberrations ( ¼ 3 independent aberrations)] and those conferring low risk [i.e., t(8;21) and t(8;21) combined with other aberrations]; Schnittger et al.86 – normal karyotype and aberrations other than t(15;17), t(8;21), inv(16)/t(16;16), t(11q23), t(6;9), t(1;3), inv(3)/t(3;3), t(3;12), t(3;21), t(8;16) and )5/)7/7q); Abu-Duhier et al.77 and Voso et al.94 – normal karyotype and +8, +21, +22, del(7q), del(9q), abnormal 11q23 and all other structural/numerical abnormalities without additional favorable [i.e., t(8;21), t(15;17) and inv(16)] or adverse [i.e., )5, )7, del(5q), abnormal 3q and complex karyotype with 5 or more abnormalities] cytogenetic changes; Moreno et al.89 – normal karyotype and aberrations other than )5/del(5q), )7/del(7q), abn 3q, complex aberrations ( P 3 independent aberrations), t(9;22), t(6;9); t(8;21) and inv(16); Rombouts et al.78 – normal karyotype and aberrations other than t(15;17), t(8;21), inv(16), t(6;9), alterations in the 11q23 region, del(5), del(7) or deletions of the q-arms of these chromosomes, and complex karyotype with more than 3 abnormalities; Kioyi et al.76 – normal karyotype and aberrations other than t(8;21), inv(16), t(9;22), 11q23 alterations, del(5) or del(7). d Intermediate-risk prognostic group included 123 patients, but numbers of patients with and without FLT3 ITD and N-RAS point mutation were not provided.
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rent AML-associated cryptic translocation, t(5;11)(q35;p15.5), was recently discovered by Brown et al.71 who used a 12-color FISH assay (termed M-TEL) capable of detecting subtle subtelomeric rearrangements that are below the resolution of G-banding. The t(5;11), which results in the NUP98-NSD1 gene fusion, was found in 2 of the 61 (3%) patients with a normal karyotype.71 The presence of an exclusively normal karyotype in a diagnostic marrow sample does not mean that cytogenetically normal leukemic blasts harbor no acquired genetic alterations. A variety of genetic abnormalities discernible only by molecular genetic techniques, such as Southern analysis, RTPCR or direct sequencing, have been reported. These include partial tandem duplication of the MLL gene (PTD MLL);29;72–75 internal tandem duplication of the FLT3 gene (FLT3 ITD);76–89 activating point mutations of D835 within the activation loop of the FLT3 gene (FLT3 Asp835 mutation);84;85;89;90 point mutations of the NRAS,76;81 CEBPA,91 and RUNX1 (AML1) genes;92 deletions and point mutations of the PU.1 gene;93 homozygous deletions resulting in null genotypes at the glutathione S-transferase GSTM1 and GSTT1 loci;94 and overexpression of the BAALC,95 and EVI1 genes.96 The molecular abnormalities shown to influence prognosis of AML patients with a normal karyotype and of those comprising the intermediate-risk cytogenetics group (which usually contains a large proportion of karyotypically normal patients) are provided in Table 3. All four studies examining the prognostic significance of the PTD MLL in patients with a normal karyotype agree that the presence of PTD MLL significantly shortens the patients’ CRD or event-free survival (EFS);72–75 in one study, OS of patients with PTD MLL was also significantly shorter than that of an age-matched karyotypically normal control group.73 The prognostic impact of FLT3 ITD, the most common gene rearrangement in AML, is more controversial. Some studies investigating patients with a normal karyotype or those comprising the intermediaterisk cytogenetics group found no significant difference in the DFS,85;86;89 EFS89 and OS80;86 between patients with and without FLT3 ITD. In other studies, patients with FLT3 ITD had significantly worse CRD,84;88 DFS,78;80 EFS86 and OS.76;77;88 These differences among studies may be attributed to several factors, including varying numbers of patients studied, differences in age of the patients, differing definitions of the intermediate-risk group (see footnote to Table 3) and thus disparate proportions of patients with particular “intermediate-risk” chromosome aberrations, and distinct induction and post-remission therapies
126
administered to patients in different studies. Additionally, some studies did not separate patients who had FLT3 ITD in combination with the FLT3 wild-type allele from those with the FLT3 ITD who lacked a FLT3 wild-type allele. Two groups have recently demonstrated that only patients with the latter genotype (or a high mutant/wt ratio, greater than 0.78) had a significantly shorter DFS and OS than patients without FLT3 ITD.80;85 Importantly, FLT3 ITD is also common in APL patients with t(15;17), but hitherto has not been shown to predict prognosis in these patients.87;88;97 This underscores the importance of assessing prognostic value of novel gene rearrangements in the context of cytogenetically defined groups of patients with AML.
Acute lymphoblastic leukemia The first comprehensive study of the cytogenetics of adult ALL was the Third International Workshop on Chromosomes in Leukemia,21 which showed that adult ALL cytogenetics have biologic and prognostic significance. Four cytogenetic classification schemata were defined: (1) according to the presence of normal and abnormal metaphases: NN – all metaphase cells normal, NN + Ncl – normal cells and non-clonal abnormalities, AN – normal cells and clonal abnormalities and AA – only abnormal metaphases; (2) according to the modal chromosome number, dividing the patients with an abnormal karyotype into six groups, 6 35, 36–45, 46 (pseudodiploid), 47–50, 51–60 and >60 chromosomes, and grouping patients with no clonal abnormalities in the “46 normal” category; (3) normal, abnormal with a translocation identified and abnormal without an identified translocation; (4) the Lund classification.21 The Lund classification was comprised of 10 hierarchical groups, t(9;22)(q34;q11.2) or a Philadelphia chromosome (Ph), t(4;11)(q21;q23), t(8;14)(q24;q32) or del(8q), other 14q+ abnormalities, del(6q), and normal karyotype, with the remaining cytogenetically abnormal patients classified by modal chromosome number as hypodiploid, pseudodiploid, low hyperdiploid (47–50 chromosomes) and high hyperdiploid (>50 chromosomes). By the Lund classification, survival was longest for patients with high hyperdiploidy and for those with normal karyotypes. Patients with t(4;11), t(8;14), 14q+, hypodiploidy and low hyperdiploidy had the shortest survival times. Remission rates were lowest in patients with hypodiploidy, t(9;22), t(4;11), t(8;14) and 14q+. Importantly, the karyotype was independent of other known prognostic factors in a
zek et al. K. Mro
multivariate analysis, although other clinical features were associated with modal number. Older age was more common in patients with low hyperdiploidy, and patients with a normal karyotype were more likely to be younger and have T-cell disease. Higher WBC was associated with pseudodiploidy, hypodiploidy, t(4;11) and t(9;22). With 6year follow-up, patients with FAB L1, WBC P50,000/ll and high or low hyperdiploidy, normal metaphases or with a del(6q) were most likely to have achieved a cure; however, the karyotype did not add prognostic significance for disease-free survival to age, WBC and immunophenotype.24 Additional reports have confirmed these observations, the clinical significance of some abnormalities has been further elucidated, and the molecular basis and biological consequences of many aberrations have been described. Below we will summarize the significance of the cytogenetics of adult ALL described since the International Workshops, particularly the more common abnormalities. Mature B-cell ALL will not be considered. The frequency of abnormal karyotypes in adult ALL is slightly higher than in pediatric ALL; 64–85%25;26;98–100 and 60–69%101;102 of successful cases, respectively. The frequencies of different ploidy groups and some aberrations, however, differ between children and adults, as does treatment outcome (Table 4). The overall survival rate for adults is 22–38%,25;99;103 whereas children have a 75–80% survival rate.104–107 The frequency of Ph+ ALL increases with age, whereas the frequency of high hyperdiploid ALL decreases with age. Less than 6% of children with ALL have a t(9;22), whereas up to 40% of adults aged P 40 years with ALL are Ph+, a poor prognostic feature regardless of age.25;108 In contrast, less than 12% of adults23;25;98;100 but 25% of children101;102;109;110 have high hyperdiploidy, a good prognostic feature. The different outcomes for adults compared with children with ALL may be partially a result of the different frequencies of specific abnormalities. A t(9;22) is the most common recurring abnormality in adult ALL, occurring in 11–29% of patients included in series comprising both B- and TALL,25;26;98 and in 37% of patients diagnosed with Bcell precursor ALL (c-ALL and pre-B ALL) who were studied by RT-PCR.103 Most (97%) Ph result from a standard t(9;22); only 3% are variant translocations. Patients with Ph+ ALL present with a high WBC (23,500/ll versus 11,550/ll),103 have B-lineage disease, a typical immunophenotype of CD34+/ CD10+/CD19+,111 and myeloid markers in up to 71% of adult cases.112 The remission rate usually is similar to the overall ALL remission rate,113 although has been lower in some studies.103 Progno-
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Table 4 Frequencies of cytogenetic aberrations in adult and childhood ALL and their prognostic relevance Abnormality
None (normal chromosomes) High hyperdiploidy Low hyperdiploidy Near-triploid/ near-tetraploid
Adults
Children
Frequencya
Prognosisa
Frequencya
Prognosisa
15–36%25;26;98–100;128
Intermediate– good26;98;127 Good25;26;98;100;127 Good25 Intermediate25
31–40%101;102
Intermediate– good101;102 Good101;102 Intermediate101;102
2–11%23;25;98;100 10–15%25;98;129 3–5%25;100 Near-triploidy: 3%98 Near-tetraploidy: 2%98
23–26%101;102;109;110 10–11%101;102 1%110;126
Near-triploidy: Poor98 Near-tetraploidy: Excellent98
Near-triploidy: Good102;126 Near-tetraploidy: Adverse126 Intermediate102 Intermediate101;102 Modal number ¼ 45: Intermediate101 Modal number ¼ 30–45: Intermediate102 Modal number <45: Poor101 Poor101;102 Poor102;171;172
Pseudodiploidy Hypodiploidy
31–50%25;98;100;129 4–9%25;98;100;129
Poor25 Poor25;98
18–26%101;102 6%101;102
Near-haploidy t(9;22)(q34;q11.2)
Not known Poor26;103;129;131;170
<1%101;102 2–6%102;108;171
t(4;11)(q21;q23) t(1;19)(q23;p13.3)
Rare100 11–29% overall25;26;98 37% of B-ALL by RT-PCR100 3–7%25;26;98;100;130;131 2–3%25;98;100;135
2%102 d 4–5%174;175
t(12;21)(p13;q22)
0–3%150–152
Poor26;133 Poor98;100;127;137 Good25 Not known
20–25%147–149
Abnormal 9p
6–30%25;26;98;100;129
Intermediate25;26;98
7–11%102;176
Abnormal 12p
4–6%25;26;98;100
7–9%102;177
del(6q)
3–16%25;26;129
6–9%102;178
Not prognostic102;178
del(7p)/del(7q)/ monosomy 7 del(5q) +8c 14q11
6–11%26;98b
Favorable25;26 Unfavorable98 Not prognostic132 Intermediate98 Adverse145 Not prognostic155 Poor26;132 Not prognostic155 Poor26 Excellent, especially t(10;14) (q24;q11)26;98
Poor173 Excellent102 Intermediate174 Good in most studies147–149 Adverse138;176 Not prognostic102 Not prognostic102;177
4%110
Adverse110
1%179 2%Unpublished data from CCG c 3–4% overall or 17–22% of T-ALL180;181
Adverse179 Unknown Not Prognostic180;181
t(10;14)(q24;q11)
<2%155 10–12%26;98 5–7% overall or 26% of T-ALL26;98 2–3%98
a
Superscript numbers denote reference numbers identifying articles that contain data on a given abnormality frequency and clinical outcome of patients with this aberration. b In Wetzler et al.26 , only patients with monosomy 7 were included. c Non-high hyperdiploid cases; CCG, Children’s Cancer Group. d Excluding infants.
sis is poor with traditional therapy and may be increasingly poor with increasing age.100 However, two studies showed a markedly reduced risk of relapse and increased survival for younger Ph+ ALL
patients who underwent allogeneic SCT from HLA-matched related donors in first CR and were conditioned with regimens that included fractionated total body irradiation and high-dose
128 etoposide.114;115 Additionally, newer biological therapy with imatinib mesylate, a drug powerfully effective in CML, another leukemia with t(9;22), also may be effective in ALL when combined with chemotherapy or other approaches. This drug competitively inhibits the abnormal protein produced in Ph+ disease,116 and may help to improve the outcome in Ph+ ALL. Early studies of Ph+ patients with ALL or CML with lymphoid blast crisis who had no response to standard treatment or who relapsed after standard treatment showed an initial 70% response rate to this therapy, but the responses have been of short duration.4 In a subsequent study of 48 patients with relapsed or refractory Ph+ ALL, imatinib induced complete hematologic and marrow responses in 29% of patients but development of resistance and subsequent disease progression were rapid, thus necessitating further studies of the efficacy of imatinib in combination with other chemotherapeutic agents in Ph+ ALL patients.116 The genes involved in the t(9;22) are ABL on chromosome 9 and BCR on chromosome 22. The 50 portion of BCR is joined to the 30 portion of ABL to form a chimeric gene, BCR-ABL. Two different breakpoints in BCR give rise to proteins of different sizes. The major breakpoint cluster region (M-bcr) spans BCR introns 12–16, and when fused with ABL results in a 210-kDa protein. This is the typical rearrangement in CML and in 24–50% of adult Ph+ ALL,113;117 but is rare in pediatric Ph+ ALL.118 The minor breakpoint (m-bcr) occurs 50 of the M-bcr in the first intron of BCR, and when fused with ABL, codes for a 190-kDa protein.119 This breakpoint occurs in 50–77% of adult Ph+ ALL103;117 and over 90% of pediatric Ph+ ALL.120 The proportion of patients with breakpoints in M-bcr increases with increasing age.121 Both proteins are tyrosine kinases, but the m-bcr product has stronger transforming potency in transfection assays and in transgenic mice.117 Comparisons of adults with Ph+ ALL with m-bcr and M-bcr breakpoints rarely show differences in the presence of additional cytogenetic aberrations, WBC, age, or outcome;117;122;123 although one study showed that patients with M-bcr breakpoints were older103 and another study of allogeneic transplants showed that patients with an m-bcr breakpoint had an increased risk of relapse compared with those with an M-bcr breakpoint.124 Cytogenetic abnormalities in addition to a t(9;22) occur in 41–86% of adult ALL at diagnosis;117;122 most frequently loss of a chromosome 7 or its short arm (7p), an additional der(22)t(9;22), abnormalities of 9p, gain of a chromosome 8 or X, duplication of 1q and high hyperdiploidy.117;122 No pretreatment clinical factors have been associated
zek et al. K. Mro with the presence of additional abnormalities.122 Outcome has been reported as no different,122;125 or dependent on the specific aberration in addition to t(9;22), with hyperdiploid patients having a better outcome and those with monosomy 7 or deletions of 9p having a particularly poor outcome.117 High hyperdiploidy is rare in adult ALL (2–11%)23;25;98;100 compared with pediatric ALL (25%).101;102;109;110 Near-triploidy and near-tetraploidy are more frequent in adult than in pediatric ALL.25;100;110;126 High hyperdiploidy in adults is associated with younger age and normal WBC. Outcome for adult high hyperdiploid patients is somewhat improved compared with other ploidy groups, but does not show the excellent outcome observed in children.25;26;98;100–102;109;110;127 Structural abnormalities are frequent in high hyperdiploid cases, most commonly a t(9;22). Patients with high hyperdiploidy and t(9;22) have outcomes similar to other Ph+ patients.98;125 Clinical features and outcome in patients with high hyperdiploidy and other recurring translocations also are similar to those of patients with the translocations and pseudodiploidy or low hyperdiploidy.25 Low hyperdiploidy is distinct from high hyperdiploidy both in chromosome pattern and in clinical features. Recent studies show an improved outcome for patients with low hyperdiploidy compared with patients with 6 46 chromosomes.25 Normal, diploid karyotypes occur in 15–36% of adult ALL25;26;98–100;128 and are associated with an intermediate26 or an improved outcome.26;98;126 Only 4–9% of adult ALL are hypodiploid.25;98;100;129 Patients with hypodiploidy are somewhat younger than those with normal karyotypes, and most have B-lineage ALL. Severe hypodiploidy with modal numbers of 30–44 is more frequent in adult ALL than is near-haploidy with chromosome numbers below 30. These patients have a poor outcome.25;98 Recurrent structural abnormalities occur in 60–70% of pseudodiploid patients,25;98 and the prognosis of these patients reflects the prognosis of the recurrent abnormality. When all pseudodiploid patients are considered, their outcome is worse than that of patients with normal chromosomes or with hyperdiploidy.25 A common recurring translocation in adult ALL is t(4;11)(q21;q23). Although particularly common in infant ALL, it also occurs in 2% of pediatric102 and 3–7% of adult ALL.25;26;98;100;130;131 It is associated with an immunophenotype of TdT+/HLA-DR+/ CD34+/CD10)/CD19+,111;132 a high WBC, a high frequency of circulating blasts and an increased frequency of central nervous system disease.132 The t(4;11) predicts a poor prognosis.26;133 Secondary abnormalities in addition to t(4;11) are
Cytogenetics in acute leukemia
common, particularly i(7)(q10) and trisomy 6; outcome does not appear to differ for these patients compared with those with t(4;11) as a sole abnormality.130 The gene involved on chromosome 11 is MLL, which has multiple translocation partners. In t(4;11), MLL is rearranged with AF4, the most common partner gene of the MLL gene in ALL. Other 11q23 (MLL) translocations also occur and overall, MLL translocations are found in up to 7% of adult ALL,26;98 occurring in both de novo and in therapy-related disease.134 Translocation (1;19)(q23;p13.3) occurs in 2–3% of adult ALL25;98;100;135 and, as in pediatric ALL, can be either balanced or an unbalanced der(19)t(1;19) with two normal chromosomes 1. Most cases are pseudodiploid, although some are high hyperdiploid. The rearrangement results in a chimeric E2APBX1 gene in 95% of cases with 6 50 chromosomes, but in only 25% of cases with >50 chromosomes.136 Nearly all cases are pre-B ALL; that is, the cells have cytoplasmic mu immunoglobulin, but not surface immunoglobulin, and are CD9+/CD10+/ CD19+/CD22+/CD34). Low WBC is common, and the patients tend to be younger.98 Outcome is poor in most studies of adults,98;100;127;137 although not all.25 Abnormalities of 9p that result in loss of genetic material [abn(9p)], including deletions, unbalanced translocations and chromosomal losses, are recurrent in ALL. Abn(9p) occurs in both T- and Blineage ALL, with some studies indicating a higher frequency in T-lineage138 and some in B-lineage ALL.139 Abn(9p) appears to be similar in adult and pediatric ALL.132 Cytogenetically, 6–30% of adult ALL patients have an abn(9p), and more show loss by molecular technologies.25;26;98;100;129 Prognosis is intermediate.25;26;98 Interestingly, several dicentric chromosomes recur in ALL; most involve chromosome arm 9p and result in partial loss of the p-arm. Dic(9;20)(p11–13;q11) has a good prognosis,140 as does dic(9;12)(p11–13;p11–12) even though most patients with the latter have at least one adverse feature.141 Dic(9;12) is more frequent in younger patients and is rare in adults.132 Abnormalities in addition to dic(9;12) are common, but complex karyotypes are rare.141 Isochromosome 9q, which results in loss of 9p, also is recurrent in ALL.142 The most common region of loss is 9p21, and deletion of the p16INK4a gene (also called CDKN2A or MTS1) appears to be the significant loss. Both hemizygous and homozygous deletions occur, with no apparent difference in clinical features or outcome.139 The neighboring gene p15INK4b (MTS2) also is lost frequently and may contribute to the pathogenesis of ALL.139 Loss of expression of p15INK4b and of p16INK4a may occur through methylation of the re-
129
maining allele in some patients with hemizygous loss.143;144 Recent studies have failed to show a prognostic significance for del(6q)132 , have shown an intermediate outcome,98 or an adverse outcome.145 Patients with del(6q) tend to be younger than those with normal 6q.145 Deletion of 6q may be a sole abnormality or part of a complex karyotype. An isolated del(6q) has been associated with hyperleukocytosis and T-ALL.145 Breakpoints vary, with band 6q21 most frequently cited as containing the smallest region of deletion.132;145 It is assumed that an as yet unidentified tumor suppressor gene is lost as a result of the deletion. Rearrangements of 12p occur in 4–6% of adult ALL,25;26;98;100 frequently as part of a complex karyotype. Translocations with many different partner chromosomes have been reported, as well as partial loss of 12p, resulting from both deletions and unbalanced translocations. Patients with 12p abnormalities tend to be younger and have a favorable prognosis in some studies,25;26 but not in all.98 One recently described aberration of 12p is the cryptic t(12;21)(p13;q22), resulting in fusion of ETV6 (TEL) and AML1 (RUNX1, CBFA2), in pre-B ALL.146 It occurs in up to 25% of children aged 1–10 years,147–149 but in only 0–3% of adult ALL.150–153 In children, it usually is associated with an excellent prognosis,147;149;151 although not always.154 Because it is infrequent in adult ALL, its prognostic significance in adults remains to be determined. Other less frequently recurring cytogenetic abnormalities in adult ALL include trisomy 8 and partial or complete monosomy for chromosomes 5 ()5/5q)) and 7 ()7/7q)).26;132;155 They are often secondary to t(9;22) and in those cases are associated with the poor outcome of Ph+ ALL.155 Patients with )5/5q) tend to be younger and frequently have T-ALL.156 Cases with )5/5q)/)7 without t(9;22) have remission and survival rates similar to patients with normal chromosomes 5 and 7,98;155 although in another study patients with )7 and +8 who were Ph-negative fared as poorly as the Ph+ patients.26 T-ALL occurs in 14–28% of adult ALL.25;26;98;100 Although the cytogenetics of T-lineage and B-lineage ALL overlap, there are distinct differences between them. In the three largest series of adult ALL,25;26;98 the proportion of cytogenetically normal cases was consistently higher in T-ALL than in B-lineage ALL (range of 24–43% versus 13–32%, respectively). About one-third of T-ALL have a translocation involving one of the T-cell receptor genes (TCR), resulting in over-expression of a gene juxtaposed to a TCR (Table 5). A breakpoint in 14q11.2, the location of a-TCR and d-TCR, is the
zek et al. K. Mro
130
Table 5 Breakpoints and genes recurrently rearranged with T-cell receptor genes in T-ALL Chromosome breakpoint
Gene involved
1p32 1p35-p34.3 8q24 9q34.3 10q24 11p13 11p15.5 19p13.2-p13.1
SCL (TAL-1 or TCL-5) LCK PVT1 and cMYC TAN-1 HOX11 RBTN2 (TTG2) RBTN1 (TTG1) LYL1
most common TCR breakpoint, but breakpoints in 7q35 (b-TCR) and 7p15 (c-TCR) also recur. The most common TCR rearrangement in adult ALL is t(10;14)(q24;q11.2), which results in overexpression of the HOX11 gene.98 It appears to be a primary aberration and is associated with a favorable outcome.98 Few studies of a recurrent d-TCR rearrangement in children, t(1;14)(p32;q11.2), have been reported in adults.31 This translocation juxtaposes d-TCR and SCL (also called TAL1 or TCL5), resulting in over-expression of SCL. A cryptic deletion of chromosome 1 in which SCL is juxtaposed to SIL also results in overexpression of SCL. In pediatric T-ALL, t(1;14) occurs only in about 3% of cases, but the SIL-SCL deletion occurs in 6–26%, making the combination of the two rearrangements the most common abnormality in pediatric TALL.157 The frequency of these rearrangements in adult ALL has not been widely studied. A cryptic t(5;14)(q35;q32) occurs in 20–30% of pediatric T-ALL;158;159 it is less common in adults. The translocation results in over-expression of HOX11L2 (5q35), although the breakpoint is actually in RanBP17, distant from HOX11L2. The abnormal expression of HOX11L2 is thought to result from abnormal control of the gene by CPT12, located at 14q32.158 Translocation (4;11)(q21;p15.5) is a rare abnormality that occurs in T-ALL,160;161 frequently in association with a del(12p). It is more frequent in children and young adults, thus far with poor outcomes. The rearrangement fuses NUP98 from chromosome 11 with RAP1GDS1 from chromosome 4 to encode nrg, a fusion protein.160;161
Concluding remarks During the last 30 years, cytogenetic analyses of patients with AML and ALL have discovered a great number of recurrent chromosome abnormalities. Several of the more common abnormalities have
been associated with specific laboratory and clinical characteristics, and are being used as diagnostic and prognostic markers that can guide the clinician in selecting the most effective treatment regimens. However, the prognostic importance of less frequent recurrent aberrations, both primary and secondary, is still unknown. Continuing cytogenetic studies are thus necessary to accrue enough patients with these rarer abnormalities to define conclusively their impact on CR rates, remission duration and survival, and to resolve discrepancies in prognostic categorization of some of the more frequent aberrations that currently exist among the major cytogenetic risk-assignment systems. Such studies will likely uncover new recurrent aberrations as they are still being identified by both classical cytogenetic methods and, increasingly, by molecular-cytogenetic techniques such as FISH, spectral karyotyping (SKY), multiplex-FISH (M-FISH) and multiplex FISH telomere assay (MTEL).38;39;68;71;158;159;162–165 Moreover, it is well known that prognostic factors depend on the type of therapy used. A cytogenetic or molecular genetic abnormality conferring an adverse prognosis with one therapeutic regimen may lose its unfavorable prognostic impact when another treatment is used. Therefore there is a constant need for large prospective studies correlating karyotype with selected molecular genetic markers, gene expression profiles, immunophenotype, other biologic parameters and clinical outcome in patients treated with both current therapies and those receiving novel therapeutic agents.
Practice points • Cytogenetic analysis should be performed in all newly diagnosed patients with AML or ALL. • Patients may be stratified to different therapies based on results of standard cytogenetic analysis, FISH and/or molecular genetic investigations (RT-PCR).
Research agenda • Continuing correlation of recurrent chromosome aberrations with response rates, response duration, survival and cure in groups of AML and ALL patients treated with current and novel induction and post-induction regimens. • Ascertainment of prognostic significance of the less common recurrent abnormalities in AML and ALL that are currently arbitrarily assigned to the intermediate-risk group or not categorized at all.
Cytogenetics in acute leukemia
• Further correlation of specific cytogenetic findings with biological and clinical features of acute leukemias to define unique disease entities that will be incorporated into the WHO Classification of Tumors of Hematopoietic and Lymphoid Tissues in the future. • Molecular genetic detection of submicroscopic mutations in patients belonging to cytogenetically defined groups (e.g., normal karyotype) that may provide additional clinically relevant information.
131
11.
12.
13.
14.
Acknowledgements 15.
This work was supported in part by grant 5P30CA16058 from the National Cancer Institute, Bethesda, MD, and the Coleman Leukemia Research Fund.
16.
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